Antibodies that immunospecifically bind to b lymphocyte stimulator protein

ABSTRACT

The present invention relates to antibodies and related molecules that immunospecifically bind to B Lymphocyte Stimulator. The present invention also relates to methods and compositions for detecting or diagnosing a disease or disorder associated with aberrant B Lymphocyte Stimulator expression or inappropriate function of B Lymphocyte Stimulator comprising antibodies or fragments or variants thereof or related molecules that immunospecifically bind to B Lymphocyte Stimulator. The present invention further relates to methods and compositions for preventing, treating or ameliorating a disease or disorder associated with aberrant B Lymphocyte Stimulator expression or inappropriate B Lymphocyte Stimulator function comprising administering to an animal an effective amount of one or more antibodies or fragments or variants thereof or related molecules that immunospecifically bind to B Lymphocyte Stimulator.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of copending U.S. patent applicationSer. No. 12/605,202, filed Oct. 23, 2009, which is continuation ofcopending U.S. patent application Ser. No. 11/054,515, filed Feb. 10,2005, which is a continuation in-part of U.S. patent application Ser.No. 10/293,418, filed Nov. 14, 2002, which issued as U.S. Pat. No.7,220,840, and U.S. patent application Ser. No. 09/880,748, filed Jun.15, 2001, which issued as U.S. Pat. No. 7,138,501. U.S. patentapplication Ser. No. 11/054,515 claims the benefit of U.S. ProvisionalApplication No. 60/543,296, filed Feb. 11, 2004, and U.S. ProvisionalApplication No. 60/580,347, filed Jun. 18, 2004. U.S. patent applicationSer. No. 10/293,418 claims the benefit of U.S. Provisional ApplicationNo. 60/331,469, filed Nov. 16, 2001, and U.S. Provisional ApplicationNo. 60/340,817, filed Dec. 19, 2001. U.S. patent application Ser. No.10/293,418 is a continuation-in-part of U.S. patent application Ser. No.09/880,748, filed Jun. 15, 2001, which issued as U.S. Pat. No.7,138,501. U.S. patent application Ser. No. 09/880,748 claims thebenefit of U.S. Provisional Application No. 60/212,210, filed Jun. 16,2000, U.S. Provisional Application No. 60/240,816, filed Oct. 17, 2000,U.S. Provisional Application No. 60/276,248, filed Mar. 16, 2001, U.S.Provisional Application No. 60/277,379, filed Mar. 21, 2001, and U.S.Provisional Application No. 60/293,499, filed May 25, 2001. Each of theapplications identified above is herein incorporated by reference intheir entireties.

INCORPORATION-BY-REFERENCE OF MATERIAL SUBMITTED ELECTRONICALLY

Incorporated by reference in its entirety herein is a computer-readablenucleotide/amino acid sequence listing submitted concurrently herewithand identified as follows: One U.S. Pat. No. 4,924,928 Byte ASCII (Text)file named “SubstituteSequenceListing.TXT,” created on Jan. 10, 2011.

INTRODUCTION

The present invention relates to antibodies and related molecules thatimmunospecifically bind to B Lymphocyte Stimulator (BLyS™) protein. Thepresent invention also relates to methods and compositions fordetecting, diagnosing, or prognosing a disease or disorder associatedwith aberrant B Lymphocyte Stimulator or B Lymphocyte Stimulatorreceptor expression or inappropriate function of B Lymphocyte Stimulatoror B Lymphocyte Stimulator receptor, comprising antibodies or fragmentsor variants thereof, or related molecules, that immunospecifically bindto B Lymphocyte Stimulator. The present invention further relates tomethods and compositions for preventing, treating or ameliorating adisease or disorder associated with aberrant B Lymphocyte Stimulator orB Lymphocyte Stimulator receptor expression or inappropriate BLymphocyte Stimulator function or B Lymphocyte Stimulator receptorfunction, comprising administering to an animal, preferably a human, aneffective amount of one or more antibodies or fragments or variantsthereof, or related molecules, that immunospecifically bind to BLymphocyte Stimulator.

BACKGROUND OF THE INVENTION

B Lymphocyte Stimulator (BLyS™) protein is a member of the tumornecrosis factor (“TNF”) superfamily that induces both in vivo and invitro B cell proliferation and differentiation (Moore et al., Science285: 260-263 (1999)). B Lymphocyte Stimulator is distinguishable fromother B cell growth and differentiation factors such as IL-2, IL-4,IL-5, IL-6, IL-7, IL-13, IL-15, CD40L, or CD27L (CD70) by itsmonocyte-specific gene and protein expression pattern and its specificreceptor distribution and biological activity on B lymphocytes. BLymphocyte Stimulator expression is not detected on natural killer(“NK”) cells, T cells or B cells, but is restricted to cells of myeloidorigin. B Lymphocyte Stimulator expression on resting monocytes isupregulated by interferon-gamma (IFN-gamma). The gene encoding BLymphocyte Stimulator has been mapped to chromosome 13q34.

B Lymphocyte Stimulator is expressed as a 285 amino acid type IImembrane-bound polypeptide and a soluble 152 amino acid polypeptide(Moore et al., 1999 supra). The membrane-bound form of B LymphocyteStimulator has a predicted transmembrane spanning domain between aminoacid residues 47 and 73. The NH₂-terminus of the soluble form of BLymphocyte Stimulator begins at Ala¹³⁴ of the membrane-bound form of BLymphocyte Stimulator. Soluble recombinant B Lymphocyte Stimulator hasbeen shown to induce in vitro proliferation of murine splenic B cellsand to bind to a cell-surface receptor on these cells (Moore et al.,1999 supra). Soluble B Lymphocyte Stimulator administration to mice hasbeen shown to result in an increase in the proportion of CD45R^(dull),Ly6D^(bright) (also known as ThB) B cells and an increase in serum IgMand IgA levels (Moore et al., 1999 supra). Thus, B Lymphocyte Stimulatordisplays a B cell tropism in both its receptor distribution andbiological activity.

Levels of B Lymphocyte Stimulator protein have been found to be elevatedin patients with autoimmune disease, including systemic lupuserythematosus (SLE), rheumatoid arthritis, and Sjögren's syndrome (Zhanget al., The Journal of Immunology, (2001) 166:6-10; Cheema et al.,Arthritis and Rheumatism (2001) 44:1313-1319; and Groom et al., Journalof Clinical Investigation (2002) 109:59-68). Furthermore, administrationof a soluble form of a B Lymphocyte Stimulator receptor, TACI, has beenshown to alleviate the autoimmune phenotype of NZBWF1 and MRL-lpr/lprmice (Gross et al., Nature, (2000) 404:995-999). Thus, antibodies andrelated molecules that immunospecifically bind to B LymphocyteStimulator may find medical utility in, for example, the treatment of Bcell disorders associated with autoimmunity In other embodiments,antibodies and related molecules that immunospecifically bind to BLymphocyte Stimulator may find medical utility in for example, neoplasiaor immunodeficiency syndromes.

SUMMARY OF THE INVENTION

The present invention encompasses antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to a polypeptide orpolypeptide fragment of B Lymphocyte Stimulator. In particular, theinvention encompasses antibodies (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof)that immunospecifically bind to a polypeptide or polypeptide fragment ofhuman B Lymphocyte Stimulator (SEQ ID NOS:3228 and/or 3229) or BLymphocyte Stimulator expressed on human monocytes; murine B LymphocyteStimulator (SEQ ID NOS:3230 and/or 3231) or B Lymphocyte Stimulatorexpressed on murine monocytes; rat B Lymphocyte Stimulator (either thesoluble forms as given in SEQ ID NOS:3232, 3233, 3234 and/or 3235 or ina membrane associated form, e.g., on the surface of rat monocytes); ormonkey B Lymphocyte Stimulator (e.g., the monkey B Lymphocyte Stimulatorpolypeptides of SEQ ID NOS:3236 and/or 3237, the soluble form of monkeyB Lymphocyte Stimulator, or B Lymphocyte Stimulator expressed on monkeymonocytes), preferably human B Lymphocyte Stimulator. The presentinvention also encompasses methods and compositions for detecting,diagnosing, or prognosing diseases or disorders associated with aberrantB Lymphocyte Stimulator or B Lymphocyte Stimulator receptor expressionor inappropriate function of B Lymphocyte Stimulator or B LymphocyteStimulator receptor in an animal, preferably a mammal, and mostpreferably a human, comprising, or alternatively consisting of, use ofantibodies (including molecules comprising, or alternatively consistingof, antibody fragments or variants thereof) that immunospecifically bindto B Lymphocyte Stimulator. Diseases and disorders which can bedetected, diagnosed, or prognosed with the antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) of the invention include, but are not limited to,immune disorders (e.g., lupus, rheumatoid arthritis, multiple sclerosis,myasthenia gravis, Hashimoto's disease, and immunodeficiency syndrome),inflammatory disorders (e.g., asthma, allergic disorders, and rheumatoidarthritis), infectious diseases (e.g., AIDS), and proliferativedisorders (e.g., leukemia, carcinoma, and lymphoma). The presentinvention further encompasses methods and compositions for preventing,treating or ameliorating diseases or disorders associated with aberrantB Lymphocyte Stimulator or B Lymphocyte Stimulator receptor expressionor inappropriate function of B Lymphocyte Stimulator or B LymphocyteStimulator receptor in an animal, preferably a mammal, and mostpreferably a human, comprising, or alternatively consisting of,administering to said animal an effective amount of one or moreantibodies (including molecules comprising, or alternatively consistingof, antibody fragments or variants thereof) that immunospecifically bindto B Lymphocyte Stimulator. Diseases and disorders which can beprevented, treated or ameliorated by administering an effective amountof an antibody of the invention include, but are not limited to, immunedisorders (e.g., lupus, rheumatoid arthritis, multiple sclerosis,myasthenia gravis, Hashimoto's disease, and immunodeficiency syndrome),inflammatory disorders (e.g., asthma, allergic disorders, and rheumatoidarthritis), infectious diseases (e.g., AIDS), and proliferativedisorders (e.g., leukemia, carcinoma, and lymphoma).

Using phage display technology, the present inventors have identifiedsingle chain antibody molecules (“scFvs”) that immunospecifically bindto B Lymphocyte Stimulator, including scFvs that immunospecifically bindto soluble B Lymphocyte Stimulator, scFvs that immunospecifically bindthe membrane-bound form of B Lymphocyte Stimulator, and scFvs thatimmunospecifically bind to both the soluble form and the membrane-boundform of B Lymphocyte Stimulator. Antibodies of the present invention aredefined as able to bind the membrane bound and/or soluble forms of BLymphocyte Stimulator according to the assays described in Examples 1through 19. Molecules comprising, or alternatively consisting of,fragments or variants of these scFvs (e.g., including VH domains, VHCDRs, VL domains, or VL CDRs having an amino acid sequence of any one ofthose referred to in Table 1), that immunospecifically bind the solubleform of B Lymphocyte Stimulator, the membrane-bound form of B LymphocyteStimulator, and/or both the soluble form and membrane-bound form of BLymphocyte Stimulator, are also encompassed by the invention, as arenucleic acid molecules that encode these scFvs, and/or molecules.

In particular, the invention relates to scFvs comprising, oralternatively consisting of, an amino acid sequence selected from thegroup consisting of SEQ ID NOS: 1-2128, preferably SEQ ID NOS:834-872,1570-1595, and 1886-1908, and most preferably SEQ ID NOS:1-46, 321-329,1563-1569, and 1881-1885, as referred to in Table 1 below. In specificembodiments, the present invention relates to scFvs thatimmunospecifically bind the soluble form of B Lymphocyte Stimulator,said scFvs comprising, or alternatively consisting of, an amino acidsequence of SEQ ID NOS: 1563-1880, preferably SEQ ID NOS:1570-1595, andmost preferably SEQ ID NOS: 1563-1569, as referred to in Table 1, below.In other embodiments, the present invention also relates to scFvs thatimmunospecifically bind the membrane-bound form of B LymphocyteStimulator, said scFvs comprising, or alternatively consisting of, anamino acid sequence of SEQ ID NOS: 1881-2128, preferably SEQ IDNOS:1886-1908, and most preferably SEQ ID NOS: 1881-1885, as referred toin Table 1 below. The present invention further relates to scFvs thatimmunospecifically bind both the membrane-bound form and soluble form ofB Lymphocyte Stimulator, said scFvs comprising, or alternativelyconsisting of, an amino acid sequence of SEQ ID NOS: 1-1562, preferablySEQ ID NOS: 834-872, and most preferably SEQ ID NOS: 1-46, and 321-329,as referred to in Table 1 below. Molecules comprising, or alternativelyconsisting of, fragments or variants of these scFvs (e.g., including VHdomains, VH CDRs, VL domains, or VL CDRs having an amino acid sequenceof any one of those referred to in Table 1), that immunospecificallybind the soluble form of B Lymphocyte Stimulator, the membrane-boundform of B Lymphocyte Stimulator, and/or both the soluble form andmembrane-bound form of B Lymphocyte Stimulator, are also encompassed bythe invention, as are nucleic acid molecules that encode these scFvs,and/or molecules.

The present invention provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to a polypeptide orpolypeptide fragment of B Lymphocyte Stimulator, said antibodiescomprising, or alternatively consisting of, a polypeptide having theamino acid sequence of any one of the variable heavy (“VH”) domainsreferred to in Table 1, below, or any one of the variable light (“VL”)domains referred to in Table 1. In a preferred embodiment, antibodies ofthe present invention comprise, or alternatively consist of, apolypeptide having the amino acid sequence of a VH domain contained inSEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908, as referredto in Table 1 below. In another preferred embodiment, antibodies(including molecules comprising or alternatively consisting of, antibodyfragments or variants thereof) of the present invention comprise, oralternatively consist of, a polypeptide having the amino acid sequenceof a VL domain contained SEQ ID NOS:1-46, 321-329, 834-872, 1563-1595,or 1881-1908, as referred to in Table 1 below. Molecules comprising, oralternatively consisting of, fragments or variants of these antibodies(e.g., including VH domains, VH CDRs, VL domains, or VL CDRs having anamino acid sequence of any one of those referred to in Table 1), thatimmunospecifically bind the soluble form of B Lymphocyte Stimulator, themembrane-bound form of B Lymphocyte Stimulator, and/or both the solubleform and membrane-bound form of B Lymphocyte Stimulator, are alsoencompassed by the invention, as are nucleic acid molecules that encodethese antibodies, and/or molecules.

The present invention also provides antibodies (including moleculescomprising or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to a polypeptide or apolypeptide fragment of B Lymphocyte Stimulator, said antibodiescomprising, or alternatively consisting of, a polypeptide having theamino acid sequence of any one of the VH domains referred to in Table 1,below, and any one of the VL domains referred to in Table 1. In apreferred embodiment, the antibodies of the invention comprise oralternatively consist of, a polypeptide having the amino acid sequenceof a VH and VL domain contained in the same scFv referred to in Table 1.In another preferred embodiment, antibodies of the present invention,comprise, or alternatively consist of, a VH domain from an scFv of SEQID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908, as disclosed inTable 1, and a VL domain from an scFv SEQ ID NOS:1-46, 321-329, 834-872,1563-1595, or 1881-1908, as disclosed in Table 1. In another preferredembodiment, antibodies of the present invention comprise, oralternatively consist of, the VH and VL domain from a single scFv of SEQID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908, as disclosed inTable 1. Molecules comprising, or alternatively consisting of, fragmentsor variants of these antibodies (e.g., including VH domains, VH CDRs, VLdomains, or VL CDRs having an amino acid sequence of any one of thosereferred to in Table 1), that immunospecifically bind the soluble formof B Lymphocyte Stimulator, the membrane-bound form of B LymphocyteStimulator, and/or both the soluble form and membrane-bound form of BLymphocyte Stimulator, are also encompassed by the invention, as arenucleic acid molecules that encode these antibodies, and/or molecules.

The present invention also provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to a polypeptide or apolypeptide fragment of B Lymphocyte Stimulator, said antibodiescomprising, or alternatively consisting of, a polypeptide having theamino acid sequence of any one, two, three or more of the VHcomplementarity determining regions (“CDRs”) (i.e., VH CDR1, VH CDR2, orVH CDR3) referred to in Table 1 and/or any one, two, three or more ofthe VL CDRs (i.e., VL CDR1, VL CDR2, or VL CDR3) referred to in Table 1.In one embodiment, antibodies of the present invention comprise, oralternatively consist of, a polypeptide having the amino acid sequenceof any one of the VH CDR1s referred to in Table 1 and/or any one of theVL CDR1s referred to in Table 1. In another embodiment, antibodies ofthe present invention comprise, or alternatively consist of, apolypeptide having the amino acid sequence of any one of the VH CDR2sreferred to in Table 1 and/or any one of the VL CDR2s referred to inTable 1. In a preferred embodiment, antibodies of the present inventioncomprise, or alternatively consist of, a polypeptide having the aminoacid sequence of any one of the VH CDR3s referred to in Table 1 and/orany one of the VL CDR3s referred to in Table 1. Molecules comprising, oralternatively consisting of, fragments or variants of these antibodies(e.g., including VH domains, VH CDRs, VL domains, or VL CDRs having anamino acid sequence of any one of those referred to in Table 1), thatimmunospecifically bind the soluble form of B Lymphocyte Stimulator, themembrane-bound form of B Lymphocyte Stimulator, and/or both the solubleform and membrane-bound form of B Lymphocyte Stimulator, are alsoencompassed by the invention, as are nucleic acid molecules that encodethese antibodies, and/or molecules.

In another embodiment, antibodies of the present invention (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) immunospecifically bind to a polypeptide orpolypeptide fragment of B Lymphocyte Stimulator, and comprise, oralternatively consist of, a polypeptide having the amino acid sequenceof any one of the VH CDR1s referred to in Table 1, any one of the VHCDR2s referred to in Table 1, and/or any one of the VH CDR3s referred toin Table 1. In another embodiment, antibodies of the present inventioncomprise, or alternatively consist of, a polypeptide having the aminoacid sequence of any one of the VL CDR1s referred to in Table 1, any oneof the VL CDR2s referred to in Table 1, and/or any one of the VL CDR3sreferred to in Table 1. In a preferred embodiment, antibodies of thepresent invention comprise, or alternatively consist of, at least one,two, three, four, five, six, or more CDRs that correspond to the samescFv referred to in Table 1, more preferably where CDR1, CDR2, and CDR3of the VL domain correspond to the same scFv or where CDR1, CDR2, andCDR3 of the VH domain correspond to the same scFv, and most preferablywhere all six CDRs correspond to the same scFv referred to in Table 1.Molecules comprising, or alternatively consisting of, fragments orvariants of these antibodies (e.g., including VH domains, VH CDRs, VLdomains, or VL CDRs having an amino acid sequence of any one of thosereferred to in Table 1), that immunospecifically bind the soluble formof B Lymphocyte Stimulator, the membrane-bound form of B LymphocyteStimulator, and/or both the soluble form and membrane-bound form of BLymphocyte Stimulator, are also encompassed by the invention, as arenucleic acid molecules that encode these antibodies, and/or molecules.

The present invention also provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to the soluble form of BLymphocyte Stimulator (e.g., a polypeptide consisting of amino acids134-285 of SEQ ID NO:3228); that immunospecifically bind to themembrane-bound form of B Lymphocyte Stimulator (e.g., a polypeptideconsisting of amino acids 1-285 of SEQ ID NO:3228 or a B LymphocyteStimulator polypeptide expressed on the surface of monocytes) and/orthat immunospecifically bind to both the soluble form and membrane-boundform of B Lymphocyte Stimulator. In a preferred embodiment, antibodiesof the present invention immunospecifically bind to the soluble form ofB Lymphocyte Stimulator and comprise, or alternatively consist of, a VHdomain, VH CDR1, VH CDR2, VH CDR3, VL domain, VL CDR1, VL CDR2, and/orVL CDR3 corresponding to one or more scFvs, that immunospecifically bindto the soluble form of B Lymphocyte Stimulator. In another preferredembodiment, antibodies of the present invention immunospecifically bindto the membrane-bound form of B Lymphocyte Stimulator and comprise, oralternatively consist of, a VH domain, VH CDR1, VH CDR2, VH CDR3, VLdomain, VL CDR1, VL CDR2, and/or VL CDR3 corresponding to one or morescFvs, that immunospecifically bind to the membrane-bound form of BLymphocyte Stimulator. In yet another preferred embodiment, antibodiesof the present invention immunospecifically bind to the soluble form andmembrane-bound form of B Lymphocyte Stimulator and comprise, oralternatively consist of, a VH domain, VH CDR1, VH CDR2, VH CDR3, VLdomain, VL CDR1, VL CDR2, and/or VL CDR3 corresponding to one or morescFvs, that immunospecifically binds to the soluble form andmembrane-bound form of B Lymphocyte Stimulator. In another preferredembodiment, antibodies of the present invention comprise, oralternatively consist of, a VH domain and a VL domain corresponding tothe same scFv disclosed in Table 1, which antibodies immunospecificallybind to the soluble form of B Lymphocyte Stimulator, the membrane-boundform of B Lymphocyte Stimulator, or both the soluble form andmembrane-bound form of B Lymphocyte Stimulator. Nucleic acid moleculesencoding these antibodies are also encompassed by the invention.Molecules comprising, or alternatively consisting of, fragments orvariants of these antibodies (e.g., including VH domains, VH CDRs, VLdomains, or VL CDRs having an amino acid sequence of any one of thosereferred to in Table 1), that immunospecifically bind the soluble formof B Lymphocyte Stimulator, the membrane-bound form of B LymphocyteStimulator, and/or both the soluble form and membrane-bound form of BLymphocyte Stimulator, are also encompassed by the invention, as arenucleic acid molecules that encode these antibodies, and/or molecules.

The present invention also provides antibodies (including moleculescomprising or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to both B LymphocyteStimulator and APRIL (preferably to the soluble forms of each of thesemolecules), said antibodies comprising, or alternatively consisting of,a polypeptide having the amino acid sequence of any one of the VHdomains referred to in Table 1, below, and any one of the VL domainsreferred to in Table 1. In a preferred embodiment, the antibodies of theinvention comprise or alternatively consist of, a polypeptide having theamino acid sequence of a VH and VL domain contained in the same scFvreferred to in Table 1. In another preferred embodiment, antibodies ofthe present invention that immunospecifically bind to both B LymphocyteStimulator and APRIL, comprise, or alternatively consist of, a VH domainfrom an scFv of SEQ ID NOS:3240-3247 as disclosed in Table 1, and a VLdomain from an scFv SEQ ID NOS:3240-3247, as disclosed in Table 1. Inanother preferred embodiment, antibodies of the present invention thatimmunospecifically bind to both B Lymphocyte Stimulator and APRILcomprise, or alternatively consist of, the VH and VL domain from asingle scFv of SEQ ID NOS: SEQ ID NOS:3240-3247, as disclosed inTable 1. Molecules comprising, or alternatively consisting of, fragmentsor variants of these antibodies (e.g., including VH domains, VH CDRs, VLdomains, or VL CDRs having an amino acid sequence of any one of thosereferred to in Table 1), that immunospecifically bind both B LymphocyteStimulator and APRIL, are also encompassed by the invention, as arenucleic acid molecules that encode these antibodies, and/or molecules.

The present invention also provides antibodies (including moleculescomprising or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to a heterotrimericprotein comprising at least one B Lymphocyte Stimulator polypeptide(preferably amino acids 134-285 of SEQ ID NO:3228), said antibodiescomprising, or alternatively consisting of, a polypeptide having theamino acid sequence of any one of the VH domains referred to in Table 1,below, and any one of the VL domains referred to in Table 1. In apreferred embodiment, the antibodies of the invention thatimmunospecifically bind heterotrimeric protein comprising at least one BLymphocyte Stimulator polypeptide, comprise or alternatively consist of,a polypeptide having the amino acid sequence of a VH and VL domaincontained in the same scFv referred to in Table 1. In another preferredembodiment, antibodies of the present invention that immunospecificallybind heterotrimeric protein comprising at least one B LymphocyteStimulator polypeptide, comprise, or alternatively consist of, a VHdomain from an scFv of SEQ ID NOS:3240-3247 as disclosed in Table 1, anda VL domain from an scFv SEQ ID NOS:3240-3247, as disclosed in Table 1.In another preferred embodiment, antibodies of the present inventionthat immunospecifically bind heterotrimeric protein comprising at leastone B Lymphocyte Stimulator polypeptide, comprise, or alternativelyconsist of, the VH and VL domain from a single scFv of SEQ IDNOS:3240-3247, as disclosed in Table 1. Molecules comprising, oralternatively consisting of, fragments or variants of these antibodies(e.g., including VH domains, VH CDRs, VL domains, or VL CDRs having anamino acid sequence of any one of those referred to in Table 1), thatimmunospecifically bind a heterotrimeric protein comprising at least oneB Lymphocyte Stimulator polypeptide, are also encompassed by theinvention, as are nucleic acid molecules that encode these antibodies,and/or molecules.

A VH domain of an amino acid sequence disclosed herein may be combinedwith a VL domain of an amino acid sequence disclosed herein, or other VLdomains, to provide a VH/VL pairing representing an antigen-binding siteof an antibody. Similarly, a VL domain of an amino acid sequencedisclosed herein may be combined with a VH domain of an amino acidsequence disclosed herein, or other VH domains. Further, one or moreCDRs disclosed herein may be taken from a VH or VL domain andincorporated into a suitable framework as discussed infra.

The present invention provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof (including derivatives)) comprising, or alternativelyconsisting of, of VH domains, VL domains and/or CDRs described herein,which antibodies, immunospecifically bind to B Lymphocyte Stimulator(e.g., soluble B Lymphocyte Stimulator and membrane-bound B LymphocyteStimulator) and can be routinely assayed for immunospecific binding to BLymphocyte Stimulator using methods known in the art, such as, forexample, the immunoassays disclosed infra. Antibodies and antibodyfragments or variants (including derivatives) of the invention mayinclude, for example, one or more amino acid sequence alterations(addition, deletion, substitution and/or insertion of an amino acidresidue). These alterations may be made in one or more framework regionsand/or one or more CDR's. The antibodies of the invention (includingantibody fragments, and variants and derivative thereof) can beroutinely made by methods known in the art. Molecules comprising, oralternatively consisting of, fragments or variants of any of the VHdomains, VH CDRs, VL domains, and VL CDRs whose sequences arespecifically disclosed herein may be employed in accordance with thepresent invention. Nucleic acid molecules encoding these antibodies andmolecules (including fragments, variants, and derivatives) are alsoencompassed by the invention.

The present invention also provides panels of antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants) wherein the panel members correspond to one, two, three,four, five, ten, fifteen, twenty, or more different antibodies of theinvention (e.g., whole antibodies, Fabs, F(ab′)₂ fragments, Fdfragments, disulfide-linked Fvs (sdFvs), antiidiotypic (anti-Id)antibodies, and scFvs). The present invention further provides mixturesof antibodies, wherein the mixture corresponds to one, two, three, four,five, ten, fifteen, twenty, or more different antibodies of theinvention (e.g., whole antibodies, Fabs, F(ab′)₂ fragments, Fdfragments, disulfide-linked Fvs (sdFvs), antiidiotypic (anti-Id)antibodies, and scFvs)). The present invention also provides forcompositions comprising, or alternatively consisting of, one, two,three, four, five, ten, fifteen, twenty, or more antibodies of thepresent invention (including molecules comprising, or alternativelyconsisting of, antibody fragments or variants thereof). A composition ofthe invention may comprise, or alternatively consist of, one, two,three, four, five, ten, fifteen, twenty, or more amino acid sequences ofone or more antibodies or fragments or variants thereof. Alternatively,a composition of the invention may comprise, or alternatively consistof, nucleic acid molecules encoding one or more antibodies of theinvention.

The present invention also provides for fusion proteins comprising anantibody (including molecules comprising, or alternatively consistingof, antibody fragments or variants thereof) of the invention, and aheterologous polypeptide (i.e., a polypeptide unrelated to an antibodyor antibody domain). Nucleic acid molecules encoding these fusionproteins are also encompassed by the invention. A composition of thepresent invention may comprise, or alternatively consist of, one, two,three, four, five, ten, fifteen, twenty or more fusion proteins of theinvention. Alternatively, a composition of the invention may comprise,or alternatively consist of, nucleic acid molecules encoding one, two,three, four, five, ten, fifteen, twenty or more fusion proteins of theinvention.

The present invention also provides for a nucleic acid molecule,generally isolated, encoding an antibody (including molecules such asscFvs, which comprise, or alternatively consist of, an antibody fragmentor variant thereof) of the invention. The present invention alsoprovides a host cell transformed with a nucleic acid molecule of theinvention and progeny thereof. The present invention also provides amethod for the production of an antibody (including a moleculecomprising, or alternatively consisting of, an antibody fragment orvariant thereof) of the invention. The present invention furtherprovides a method of expressing an antibody (including a moleculecomprising, or alternatively consisting of, an antibody fragment orvariant thereof) of the invention from a nucleic acid molecule. Theseand other aspects of the invention are described in further detailbelow.

The present invention also encompasses methods and compositions fordetecting, diagnosing and/or prognosing diseases or disorders associatedwith aberrant B Lymphocyte Stimulator or B Lymphocyte Stimulatorreceptor expression or inappropriate B Lymphocyte Stimulator or BLymphocyte Stimulator receptor function in an animal, preferably amammal, and most preferably a human, comprising using antibodies(including molecules which comprise, or alternatively consist of,antibody fragments or variants thereof) that immunospecifically bind toB Lymphocyte Stimulator. Diseases and disorders which can be detected,diagnosed or prognosed with the antibodies of the invention include, butare not limited to, immune disorders (e.g., lupus, rheumatoid arthritis,multiple sclerosis, myasthenia gravis, Hashimoto's disease, andimmunodeficiency syndrome), inflammatory disorders (e.g., asthma,allergic disorders, and rheumatoid arthritis), infectious diseases(e.g., AIDS), and proliferative disorders (e.g., leukemia, carcinoma,and lymphoma).

In specific embodiments, the present invention encompasses methods andcompositions for detecting, diagnosing and/or prognosing diseases ordisorders associated with hypergammaglobulinemia (e.g., AIDS, autoimmunediseases, and some immunodeficiencies). In other specific embodiments,the present invention encompasses methods and compositions fordetecting, diagnosing and/or prognosing diseases or disorders associatedwith hypogammaglobulinemia (e.g., an immunodeficiency).

The present invention further encompasses methods and compositions forpreventing, treating or ameliorating diseases or disorders associatedwith aberrant B Lymphocyte Stimulator or B Lymphocyte Stimulatorreceptor expression or inappropriate B Lymphocyte Stimulator or BLymphocyte Stimulator receptor function in an animal, preferably amammal, and most preferably a human, comprising administering to saidanimal an effective amount of one or more antibodies (includingmolecules which comprise, or alternatively consist of, antibodyfragments or variants thereof) that immunospecifically bind to BLymphocyte Stimulator. Diseases and disorders which can be prevented,treated or inhibited by administering an effective amount of one or moreantibodies or molecules of the invention include, but are not limitedto, immune disorders (e.g., lupus, rheumatoid arthritis, multiplesclerosis, myasthenia gravis, Hashimoto's disease, and immunodeficiencysyndrome), inflammatory disorders (e.g., asthma, allergic disorders, andrheumatoid arthritis), infectious diseases (e.g., AIDS), andproliferative disorders (e.g., leukemia, carcinoma, and lymphoma).

In specific embodiments, the present invention encompasses methods andcompositions (e.g., antagonistic anti-B Lymphocyte Stimulatorantibodies) for preventing, treating or ameliorating diseases ordisorders associated with hypergammaglobulinemia (e.g., AIDS, autoimmunediseases, and some immunodeficiency syndromes). In other specificembodiments, the present invention encompasses methods and compositions(e.g., agonistic anti-B Lymphocyte Stimulator antibodies) forpreventing, treating or ameliorating diseases or disorders associatedwith hypogammaglobulinemia (e.g., an immunodeficiency syndrome).

Autoimmune disorders, diseases, or conditions that may be detected,diagnosed, prognosed, or monitored using the antibodies of the inventioninclude, but are not limited to, autoimmune hemolytic anemia, autoimmuneneonatal thrombocytopenia, idiopathic thrombocytopenia purpura,autoimmune neutropenia, autoimmunocytopenia, hemolytic anemia,antiphospholipid syndrome, dermatitis, gluten-sensitive enteropathy,allergic encephalomyelitis, myocarditis, relapsing polychondritis,rheumatic heart disease, glomerulonephritis (e.g., IgA nephropathy),Multiple Sclerosis, Neuritis, Uveitis Ophthalmia, Polyendocrinopathies,Purpura (e.g., Henloch-Scoenlein purpura), Reiter's Disease, Stiff-ManSyndrome, Autoimmune Pulmonary Inflammation, myocarditis, IgAglomerulonephritis, dense deposit disease, rheumatic heart disease,Guillain-Barre Syndrome, insulin dependent diabetes mellitis, andautoimmune inflammatory eye, autoimmune thyroiditis, hypothyroidism(i.e., Hashimoto's thyroiditis, systemic lupus erhythematosus, discoidlupus, Goodpasture's syndrome, Pemphigus, Receptor autoimmunities suchas, for example, (a) Graves' Disease, (b) Myasthenia Gravis, and (c)insulin resistance, autoimmune hemolytic anemia, autoimmunethrombocytopenic purpura, rheumatoid arthritis, schleroderma withanti-collagen antibodies, mixed connective tissue disease,polymyositis/dermatomyositis, pernicious anemia, idiopathic Addison'sdisease, infertility, glomerulonephritis such as primaryglomerulonephritis and IgA nephropathy, bullous pemphigoid, Sjögren'ssyndrome, diabetes mellitus, and adrenergic drug resistance (includingadrenergic drug resistance with asthma or cystic fibrosis), chronicactive hepatitis, primary biliary cirrhosis, other endocrine glandfailure, vitiligo, vasculitis, post-MI, cardiotomy syndrome, urticaria,atopic dermatitis, asthma, inflammatory myopathies, and otherinflammatory, granulomatous, degenerative, and atrophic disorders).

Immunodeficiencies that may be detected, diagnosed, prognosed, ormonitored using the antibodies of the invention include, but are notlimited to, severe combined immunodeficiency (SCID)-X linked,SCID-autosomal, adenosine deaminase deficiency (ADA deficiency),X-linked agammaglobulinemia (XLA), Bruton's disease, congenitalagammaglobulinemia, X-linked infantile agammaglobulinemia, acquiredagammaglobulinemia, adult onset agammaglobulinemia, late-onsetagammaglobulinemia, dysgammaglobulinemia, hypogammaglobulinemia,transient hypogammaglobulinemia of infancy, unspecifiedhypogammaglobulinemia, agammaglobulinemia, common variableimmunodeficiency (CVID) (acquired), Wiskott-Aldrich Syndrome (WAS),X-linked immunodeficiency with hyper IgM, non X-linked immunodeficiencywith hyper IgM, selective IgA deficiency, IgG subclass deficiency (withor without IgA deficiency), antibody deficiency with normal or elevatedIgs, immunodeficiency with thymoma, Ig heavy chain deletions, kappachain deficiency, B cell lymphoproliferative disorder (BLPD), selectiveIgM immunodeficiency, recessive agammaglobulinemia (Swiss type),reticular dysgenesis, neonatal neutropenia, severe congenitalleukopenia, thymic alymphoplasia-aplasia or dysplasia withimmunodeficiency, ataxia-telangiectasia, short limbed dwarfism, X-linkedlymphoproliferative syndrome (XLP), Nezelof syndrome-combinedimmunodeficiency with Igs, purine nucleoside phosphorylase deficiency(PNP), MHC Class II deficiency (Bare Lymphocyte Syndrome) and severecombined immunodeficiency.

Definitions

The term “antibody,” as used herein, refers to immunoglobulin moleculesand immunologically active portions of immunoglobulin molecules, i.e.,molecules that contain an antigen binding site that immunospecificallybinds an antigen. As such, the term antibody encompasses not only wholeantibody molecules, but also antibody fragments as well as variants(including derivatives) of antibodies and antibody fragments. Examplesof molecules which are described by the term “antibody” in thisapplication include, but are not limited to: single chain Fvs (scFvs),Fab fragments, Fab′ fragments, F(ab′)₂, disulfide linked Fvs (sdFvs),Fvs, and fragments comprising or alternatively consisting of, either aVL or a VH domain. The term “single chain Fv” or “scFv” as used hereinrefers to a polypeptide comprising a VL domain of antibody linked to aVH domain of an antibody. Antibodies that immunospecifically bind to BLymphocyte Stimulator may have cross-reactivity with other antigens.Preferably, antibodies that immunospecifically bind to B LymphocyteStimulator do not cross-react with other antigens. Antibodies thatimmunospecifically bind to B Lymphocyte Stimulator can be identified,for example, by immunoassays or other techniques known to those of skillin the art, e.g., the immunoassays described in the Examples below.

Antibodies of the invention include, but are not limited to, monoclonal,multispecific, human or chimeric antibodies, single chain antibodies,Fab fragments, F(ab′) fragments, anti-idiotypic (anti-Id) antibodies(including, e.g., anti-Id antibodies to antibodies of the invention),and epitope-binding fragments of any of the above. The immunoglobulinmolecules of the invention can be of any type (e.g., IgG, IgE, IgM, IgD,IgA and IgY), class (e.g., IgG₁, IgG₂, IgG₃, IgG₄, IgA₁ and IgA₂) orsubclass of immunoglobulin molecule.

Antibodies of the invention may also include multimeric forms ofantibodies. For example, antibodies of the invention may take the formof antibody dimers, trimers, or higher-order multimers of monomericimmunoglobulin molecules. Dimers of whole immunoglobulin molecules or ofF(ab′)₂ fragments are tetravalent, whereas dimers of Fab fragments orscFv molecules are bivalent. Individual monomers withon an antibodymultimer may be identical or different, i.e., they may be heteromeric orhomomeric antibody multimers. For example, individual antibodies withina multimer may have the same or different binding specificities.Multimerization of antibodies may be accomplished through naturalaggregation of antibodies or through chemical or recombinant linkingtechniques known in the art. For example, some percentage of purifiedantibody preparations (e.g., purified IgG1 molecules) spontaneously formprotein aggregates containing antibody homodimers, and otherhigher-order antibody multimers. Alternatively, antibody homodimers maybe formed through chemical linkage techniques known in the art. Forexample, heterobifunctional crosslinking agents including, but notlimited to, SMCC [succinimidyl4-(maleimidomethyl)cyclohexane-1-carboxylate] and SATA [N-succinimidylS-acethylthio-acetate] (available, for example, from PierceBiotechnology, Inc. (Rockford, Ill.)) can be used to form antibodymultimers. An exemplary protocol for the formation of antibodyhomodimers is given in Ghetie et al., Proceedings of the NationalAcademy of Sciences USA (1997) 94:7509-7514, which is herebyincorporated by reference in its entirety. Antibody homodimers can beconverted to Fab′2 homodimers through digestion with pepsin. Another wayto form antibody homodimers is through the use of the autophilic T15peptide described in Zhao and Kohler, The Journal of Immunology (2002)25:396-404, which is hereby incorporated by reference in its entirety.

Alternatively, antibodies can be made to multimerize through recombinantDNA techniques. IgM and IgA naturally form antibody multimers throughthe interaction with the J chain polypeptide. Non-IgA or non-IgMmolecules, such as IgG molecules, can be engineered to contain the Jchain interaction domain of IgA or IgM, thereby conferring the abilityto form higher order multimers on the non-IgA or non-IgM molecules.(see, for example, Chintalacharuvu et al., (2001) Clinical Immunology101:21-31. and Frigerio et al., (2000) Plant Physiology 123:1483-94,both of which are hereby incorporated by reference in their entireties.)ScFv dimers can also be formed through recombinant techniques known inthe art; an example of the construction of scFv dimers is given in Goelet al., (2000) Cancer Research 60:6964-6971 which is hereby incorporatedby reference in its entirety. Antibody multimers may be purified usingany suitable method known in the art, including, but not limited to,size exclusion chromatography.

Unless otherwise defined in the specification, specific binding orimmunospecifc binding by an anti-B Lymphocyte Stimulator antibody meansthat the anti-B Lymphocyte Stimulator antibody binds B LymphocyteStimulator but does not significantly bind to (i.e., cross react with)proteins other than B Lymphocyte Stimulator, such as other proteins inthe same family of proteins, e.g., other TNF family ligands). Anantibody that binds B Lymphocyte Stimulator protein and does notcross-react with other proteins is not necessarily an antibody that doesnot bind said other proteins in all conditions; rather, the B LymphocyteStimulator -specific antibody of the invention preferentially binds BLymphocyte Stimulator compared to its ability to bind said otherproteins such that it will be suitable for use in at least one type ofassay or treatment, i.e., give low background levels or result in nounreasonable adverse effects in treatment. It is well known that theportion of a protein bound by an antibody is known as the epitope. Anepitope may either be linear (i.e., comprised of sequential amino acidsresidues in a protein sequences) or conformational (i.e., comprised ofone or more amino acid residues that are not contiguous in the primarystructure of the protein but that are brought together by the secondary,tertiary or quaternary structure of a protein). Given that B LymphocyteStimulator -specific antibodies bind to epitopes of B LymphocyteStimulator, an antibody that specifically binds B Lymphocyte Stimulatormay or may not bind fragments of B Lymphocyte Stimulator and/or variantsof B Lymphocyte Stimulator (e.g., proteins that are at least 90%identical to B Lymphocyte Stimulator) depending on the presence orabsence of the epitope bound by a given B Lymphocyte Stimulator-specificantibody in the B Lymphocyte Stimulator fragment or variant. Likewise, BLymphocyte Stimulator-specific antibodies of the invention may bindspecies orthologues of B Lymphocyte Stimulator (including fragmentsthereof) depending on the presence or absence of the epitope recognizedby the antibody in the orthologue. Additionally, B LymphocyteStimulator-specific antibodies of the invention may bind modified formsof B Lymphocyte Stimulator, for example, B Lymphocyte Stimulator fusionproteins. In such a case when antibodies of the invention bind BLymphocyte Stimulator fusion proteins, the antibody must make bindingcontact with the B Lymphocyte Stimulator moiety of the fusion protein inorder for the binding to be specific. Antibodies that specifically bindto B Lymphocyte Stimulator can be identified, for example, byimmunoassays or other techniques known to those of skill in the art,e.g., the immunoassays described in the Examples below.

Furthermore, in the present application certain antibodies may bespecific for either the membrane bound form of B Lymphocyte Stimulator,or the soluble form of B Lymphocyte Stimulator (i.e., 134-285 of SEQ IDNO:2, preferably trimers of proteins consisting of amino acids 134-285of SEQ ID NO:2), or both. Antibodies of the present invention aredefined as able to bind the membrane bound and/or soluble forms of BLymphocyte Stimulator according to the assays described in Examples 1through 19.

Preferably, an antibody of the invention comprises, or alternativelyconsists of, a VH domain, VH CDR, VL domain, or VL CDR having an aminoacid sequence of any one of those referred to in Table 1, or a fragmentor variant thereof.

An antibody of the invention “which binds the soluble form of BLymphocyte Stimulator” is one which binds the 152 amino acid solubleform of the B Lymphocyte Stimulator protein (amino acids 134-285 of SEQID NO:3228). In specific embodiments of the invention, an antibody ofthe invention “which binds the soluble form of B Lymphocyte Stimulator”does not also bind the membrane-bound or membrane-associated form of BLymphocyte Stimulator. Assays which measure binding to the soluble formof B Lymphocyte Stimulator include, but are not limited to, receptorbinding inhibition assay or capture of soluble B Lymphocyte Stimulatorfrom solution as described in Examples 8 and 9.

An antibody of the invention “which binds the membrane-bound form of BLymphocyte Stimulator” is one which binds the membrane-associated(uncleaved) B Lymphocyte Stimulator protein. In specific embodiments ofthe invention, an antibody of the invention “which binds themembrane-bound form of B Lymphocyte Stimulator” does not also bind thesoluble form of B Lymphocyte Stimulator. Binding to HIS-tagged BLymphocyte Stimulator (as described herein) in an ELISA is an indicatorthat an antibody binds the membrane-bound form of B LymphocyteStimulator, but should not be relied upon as proof of specificity forthe membrane-bound form of B Lymphocyte Stimulator. Assays that may berelied upon as proof of an antibody's specificity for membrane-bound BLymphocyte Stimulator, include, but are not limited to, binding toplasma membranes expressing B Lymphocyte Stimulator as described inExample 2. An antibody of the invention “which binds the both thesoluble form and the membrane-bound form of B Lymphocyte Stimulator” isone which binds both the membrane-bound form and the soluble form of BLymphocyte Stimulator.

The term “variant” as used herein refers to a polypeptide that possessesa similar or identical function as a B Lymphocyte Stimulatorpolypeptide, a fragment of B Lymphocyte Stimulator, an anti-B LymphocyteStimulator antibody or antibody fragment thereof, but does notnecessarily comprise a similar or identical amino acid sequence of a BLymphocyte Stimulator polypeptide, a fragment of B LymphocyteStimulator, an anti-B Lymphocyte Stimulator antibody or antibodyfragment thereof, or possess a similar or identical structure of a BLymphocyte Stimulator polypeptide, a fragment of B LymphocyteStimulator, an anti-B Lymphocyte Stimulator antibody or antibodyfragment thereof. A variant having a similar amino acid refers to apolypeptide that satisfies at least one of the following: (a) apolypeptide comprising, or alternatively consisting of, an amino acidsequence that is at least 30%, at least 35%, at least 40%, at least 45%,at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, atleast 75%, at least 80%, at least 85%, at least 90%, at least 95% or atleast 99% identical to the amino acid sequence of a B LymphocyteStimulator polypeptide, a fragment of B Lymphocyte Stimulator, an anti-BLymphocyte Stimulator antibody or antibody fragment thereof (including aVH domain, VHCDR, VL domain, or VLCDR having an amino acid sequence ofany one of those referred to in Table 1) described herein; (b) apolypeptide encoded by a nucleotide sequence, the complementary sequenceof which hybridizes under stringent conditions to a nucleotide sequenceencoding a B Lymphocyte Stimulator polypeptide (e.g., SEQ ID NO:3228), afragment of B Lymphocyte Stimulator, an anti-B Lymphocyte Stimulatorantibody or antibody fragment thereof (including a VH domain, VHCDR, VLdomain, or VLCDR having an amino acid sequence of any one of thosereferred to in Table 1), described herein, of at least 5 amino acidresidues, at least 10 amino acid residues, at least 15 amino acidresidues, at least 20 amino acid residues, at least 25 amino acidresidues, at least 30 amino acid residues, at least 40 amino acidresidues, at least 50 amino acid residues, at least 60 amino residues,at least 70 amino acid residues, at least 80 amino acid residues, atleast 90 amino acid residues, at least 100 amino acid residues, at least125 amino acid residues, or at least 150 amino acid residues; and (c) apolypeptide encoded by a nucleotide sequence that is at least 30%, atleast 35%, at least 40%, at least 45%, at least 50%, at least 55%, atleast 60%, at least 65%, at least 70%, at least 75%, at least 80%, atleast 85%, at least 90%, at least 95% or at least 99%, identical to thenucleotide sequence encoding a B Lymphocyte Stimulator polypeptide, afragment of B Lymphocyte Stimulator, an anti-B Lymphocyte Stimulatorantibody or antibody fragment thereof (including a VH domain, VHCDR, VLdomain, or VLCDR having an amino acid sequence of any one of thosereferred to in Table 1), described herein. A polypeptide with similarstructure to a B Lymphocyte Stimulator polypeptide, a fragment of BLymphocyte Stimulator, an anti-B Lymphocyte Stimulator antibody orantibody fragment thereof, described herein refers to a polypeptide thathas a similar secondary, tertiary or quarternary structure of a BLymphocyte Stimulator polypeptide, a fragment of B LymphocyteStimulator, an anti-B Lymphocyte Stimulator antibody, or antibodyfragment thereof, described herein. The structure of a polypeptide candetermined by methods known to those skilled in the art, including butnot limited to, X-ray crystallography, nuclear magnetic resonance, andcrystallographic electron microscopy.

To determine the percent identity of two amino acid sequences or of twonucleic acid sequences, the sequences are aligned for optimal comparisonpurposes (e.g., gaps can be introduced in the sequence of a first aminoacid or nucleic acid sequence for optimal alignment with a second aminoacid or nucleic acid sequence). The amino acid residues or nucleotidesat corresponding amino acid positions or nucleotide positions are thencompared. When a position in the first sequence is occupied by the sameamino acid residue or nucleotide at the corresponding position in thesecond sequence, then the molecules are identical at that position. Thepercent identity between the two sequences is a function of the numberof identical positions shared by the sequences (i.e., % identity=numberof identical overlapping positions/total number of positions×100%). Inone embodiment, the two sequences are the same length.

The determination of percent identity between two sequences can beaccomplished using a mathematical algorithm known to those of skill inthe art. An example of a mathematical algorithm for comparing twosequences is the algorithm of Karlin and Altschul Proc. Natl. Acad. Sci.USA 87:2264-2268(1990), modified as in Karlin and Altschul Proc. Natl.Acad. Sci. USA 90:5873-5877(1993). The BLASTn and BLASTx programs ofAltschul, et al. J. Mol. Biol. 215:403-410(1990) have incorporated suchan alogrithm. BLAST nucleotide searches can be performed with the BLASTnprogram, score=100, wordlength=12 to obtain nucleotide sequenceshomologous to a nucleic acid molecules of the invention. BLAST proteinsearches can be performed with the BLASTx program, score=50,wordlength=3 to obtain amino acid sequences homologous to a proteinmolecules of the invention. To obtain gapped alignments for comparisonpurposes, Gapped BLAST can be utilized as described in Altschul et al.Nucleic Acids Res. 25:3389-3402(1997). Alternatively, PSI-BLAST can beused to perform an iterated search which detects distant relationshipsbetween molecules (Id.). When utilizing BLAST, Gapped BLAST, andPSI-BLAST programs, the default parameters of the respective programs(e.g., BLASTx and BLASTn) can be used. (Seehttp://www.ncbi.nlm.nih.gov.)

Another example of a mathematical algorithm utilized for the comparisonof sequences is the algorithm of Myers and Miller, CABIOS (1989). TheALIGN program (version 2.0) which is part of the GCG sequence alignmentsoftware package has incorporated such an alogrithm. Other algorithmsfor sequence analysis known in the art include ADVANCE and ADAM asdescribed in Torellis and Robotti Comput. Appl. Biosci., 10 :3-5(1994);and FASTA described in Pearson and Lipman Proc. Natl. Acad. Sci.85:2444-8(1988). Within FASTA, ktup is a control option that sets thesensitivity and speed of the search.

The term “derivative” as used herein, refers to a variant polypeptide ofthe invention that comprises, or alternatively consists of, an aminoacid sequence of a B Lymphocyte Stimulator polypeptide, a fragment of BLymphocyte Stimulator, or an antibody of the invention thatimmunospecifically binds to B Lymphocyte Stimulator, which has beenaltered by the introduction of amino acid residue substitutions,deletions or additions. The term “derivative” as used herein also refersto a B Lymphocyte Stimulator polypeptide, a fragment of B LymphocyteStimulator, an antibody that immunospecifically binds to B LymphocyteStimulator which has been modified, e.g., by the covalent attachment ofany type of molecule to the polypeptide. For example, but not by way oflimitation, a B Lymphocyte Stimulator polypeptide, a fragment of BLymphocyte Stimulator, or an anti-B Lymphocyte Stimulator antibody, maybe modified, e.g., by glycosylation, acetylation, pegylation,phosphorylation, amidation, derivatization by known protecting/blockinggroups, proteolytic cleavage, linkage to a cellular ligand or otherprotein, etc. A derivative of a B Lymphocyte Stimulator polypeptide, afragment of B Lymphocyte Stimulator, or an anti-B Lymphocyte Stimulatorantibody, may be modified by chemical modifications using techniquesknown to those of skill in the art, including, but not limited to,specific chemical cleavage, acetylation, formylation, metabolicsynthesis of tunicamycin, etc. Further, a derivative of a B LymphocyteStimulator polypeptide, a fragment of B Lymphocyte Stimulator, or ananti-B Lymphocyte Stimulator antibody, may contain one or morenon-classical amino acids. A polypeptide derivative possesses a similaror identical function as a B Lymphocyte Stimulator polypeptide, afragment of B Lymphocyte Stimulator, or an anti-B Lymphocyte Stimulatorantibody, described herein.

The term “epitopes” as used herein refers to portions of B LymphocyteStimulator having antigenic or immunogenic activity in an animal,preferably a mammal An epitope having immunogenic activity is a portionof B Lymphocyte Stimulator that elicits an antibody response in ananimal. An eptiope having antigenic activity is a portion of BLymphocyte Stimulator to which an antibody immunospecifically binds asdetermined by any method known in the art, for example, by theimmunoassays described herein. Antigenic epitopes need not necessarilybe immunogenic.

The term “fragment” as used herein refers to a polypeptide comprising anamino acid sequence of at least 5 amino acid residues, at least 10 aminoacid residues, at least 15 amino acid residues, at least 20 amino acidresidues, at least 25 amino acid residues, at least 30 amino acidresidues, at least 35 amino acid residues, at least 40 amino acidresidues, at least 45 amino acid residues, at least 50 amino acidresidues, at least 60 amino residues, at least 70 amino acid residues,at least 80 amino acid residues, at least 90 amino acid residues, atleast 100 amino acid residues, at least 125 amino acid residues, atleast 150 amino acid residues, at least 175 amino acid residues, atleast 200 amino acid residues, or at least 250 amino acid residues, ofthe amino acid sequence of B Lymphocyte Stimulator, or an anti-BLymphocyte Stimulator antibody (including molecules such as scFv's, thatcomprise, or alternatively consist of, antibody fragments or variantsthereof) that immunospecifically binds to B Lymphocyte Stimulator.

The term “fusion protein” as used herein refers to a polypeptide thatcomprises, or alternatively consists of, an amino acid sequence of ananti-B Lymphocyte Stimulator antibody of the invention and an amino acidsequence of a heterologous polypeptide (i.e., a polypeptide unrelated toan antibody or antibody domain).

The term “host cell” as used herein refers to the particular subjectcell transfected with a nucleic acid molecule and the progeny orpotential progeny of such a cell. Progeny may not be identical to theparent cell transfected with the nucleic acid molecule due to mutationsor environmental influences that may occur in succeeding generations orintegration of the nucleic acid molecule into the host cell genome.

By “isolated antibody” is intended an antibody removed from its nativeenvironment. Thus, an antibody produced and/or contained within arecombinant host cell is considered isolated for purposes of the presentinvention.

DESCRIPTION OF THE FIGURES

FIG. 1. ELISA results for three scFvs, I006E07, I008D05 and I016F04,that immunospecifically bind to U937 membranes, but not to bind to orcross-react with TNF-alpha or BSA.

FIG. 2. The results for three scFvs, I016H07, I001C09 and I018D07, in areceptor inhibition assay.

FIG. 3. ELISA results for two scFvs (I022D01 and I031F02) demonstratingtheir ability to bind to human B Lymphocyte Stimulator and tocross-react with mouse B Lymphocyte Stimulator, but not to bind to orcross-react with other antigens of the TNF ligand family.

FIG. 4. ELISA results for three scFvs (I031F09, I050A12, and I051C04)binding to U937 plasma membranes when either B Lymphocyte Stimulator orTNF-alpha is used as a competitor.

FIG. 5. Kinetic analysis of scFv antibody I003CO2. A dilution series ofI003CO2 from 3 nM to 825 nM is shown. Association and dissociationcurves were generated using a BIAcore 2000 and BIAevaluation 3.0software.

FIG. 6. Typical titration curves for two scFv antibodies (I007F11 andI050A07) are shown in FIG. 6. Unlabelled B Lymphocyte Stimulatorcompeted for binding to its receptor with an IC₅₀ value of 0.8 nM. TheIC₅₀ values for I007F11 and 1050A07 are 7.9 nM and 17.1 nM,respectively. The assay was performed in triplicate and standard errorbars are shown.

FIG. 7. ELISA results for three scFvs clones (I074B12, I075F12 andI075A02) that immunospecifically bind to immobilized B LymphocyteStimulator, but not to U937 plasma membranes, TNF-alpha or BSA. As acontrol, a phage antibody that recognizes TNFα, is also shown in FIG. 7.

FIG. 8. The results for two scFvs (I025B09 and I026C04) in a receptorinhibition assay.

FIG. 9. ELISA results for two scFvs clones (I067F05 and I078D02)demonstrating their ability to bind to immobilized human B LymphocyteStimulator and to cross-react with immobilized mouse B LymphocyteStimulator, but not to bind to or cross-react with other antigens of theTNF ligand family.

As a control, a phage antibody that recognizes TNFα, is also shown inFIG. 7.

FIG. 10. Kinetic analysis of scFV antibody I002A01. A dilution series ofI002A01from 3 nM to 1650 nM is shown. Association and dissociationcurves were generated using a BIAcore 2000 and BIAevaluation 3.0software.

FIG. 11. Typical titration curves for two scFvs, I0068C06 and I074B12,are shown in FIG. 11. Unlabelled B Lymphocyte Stimulator competed forbinding to its receptor with an inhibitory constant 50 (IC₅₀) value of0.66 nM. The IC₅₀ values for 0068C06 and I074B12 are 62 nM and 13 nM,respectively. The assay was performed in triplicate and standard errorbars are shown.

FIG. 12. ELISA results for three clones (I079C01, I081C10 and I082A02)demonstrating their ability to bind histidine-tagged B LymphocyteStimulator, U937 plasma membranes, but not to bind immobilizedbiotinylated B Lymphocyte Stimulator.

FIG. 13. ELISA results for three scFvs (I079B04, I079F08, and I080B01)binding to U937 plasma membranes when either histidine-tagged BLymphocyte Stimulator or biotinylated B Lymphocyte Stimulator is used asa competitor.

FIG. 14. An example of the dissociation section of a typical sensorgramfor 8 scFvs is shown in FIG. 14. An anti-TNFα antibody that does notrecognize B Lymphocyte Stimulator was included as a control. Of the 8scFvs exemplified, I079F06 was identified for further study due to therelatively high numbers of RU's bound to the surface.

FIG. 15. A typical example of the binding curves generated for the scFvantibody I082CO3 is shown in FIG. 15. The off-rate for this clone wascalculated as 2×10⁻³ s⁻¹. The affinity of I082CO3 was calculated as 20nM, assuming 100% activity of the scFv.

FIG. 16. ELISA results for three scFvs (I079B04, I079F08, and I080B01)binding to P388 plasma membranes when either histidine-tagged BLymphocyte Stimulator or biotinylated B Lymphocyte Stimulator is used asa competitor.

DETAILED DESCRIPTION OF THE INVENTION

The present invention encompasses antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to B LymphocyteStimulator or a fragment or variant of B Lymphocyte Stimulator. Inparticular, the invention provides antibodies such as, for example,single chain Fvs (scFvs) having an amino acid sequence of any one of SEQID NOS:1-2128, as referred to in Table 1. In particular, the presentinvention encompasses antibodies that immunospecifically bind to apolypeptide, a polypeptide fragment or variant, or an epitope of human BLymphocyte Stimulator (SEQ ID NOS:3228 and/or 3229) or B LymphocyteStimulator expressed on human monocytes; murine B Lymphocyte Stimulator(SEQ ID NOS:3230 and/or 3231) or B Lymphocyte Stimulator expressed onmurine monocytes; rat B Lymphocyte Stimulator (either the soluble formsas given in SEQ ID NOS:3232, 3233, 3234 and/or 3235 or in a membraneassociated form, e.g., on the surface of rat monocytes); or monkey BLymphocyte Stimulator (e.g., the monkey B Lymphocyte Stimulatorpolypeptides of SEQ ID NOS:3236 and/or 3237, the soluble form of monkeyB Lymphocyte Stimulator, or B Lymphocyte Stimulator expressed on monkeymonocytes) (as determined by immunoassays known in the art for assayingspecific antibody-antigen binding).

The polypeptide sequence shown in SEQ ID NO:3228 was obtained bysequencing and translating the cDNA of the HNEDU15 clone which wasdeposited on Oct. 22, 1996 at the American Type Culture Collection,10801 University Boulevard, Manassas, Va. 20110-2209, and assigned ATCC™Accession No. 97768. The deposited clone is contained in the pBluescriptSK(−) plasmid (Stratagene, La Jolla, Calif.). The ATCC™ deposits weremade pursuant to the terms of the Budapest Treaty on the internationalrecognition of the deposit of microorganisms for the purposes of patentprocedure.

The polypeptide sequence shown in SEQ ID NO:3229 was obtained bysequencing and translating the cDNA of the HDPMC52 clone, which wasdeposited on Dec. 10, 1998 at the American Type Culture Collection, andassigned ATCC™ Accession No. 203518. The deposited clone is contained inthe pBluescript SK(−) plasmid (Stratagene, La Jolla, Calif.). The ATCC™deposits were made pursuant to the terms of the Budapest Treaty on theinternational recognition of the deposit of microorganisms for thepurposes of patent procedure.

The B Lymphocyte Stimulator polypeptides bound by the antibodies of theinvention may be in monomers or multimers (i.e., dimers, trimers,tetramers and higher multimers). Accordingly, the present inventionrelates to antibodies that bind monomers and multimers of the BLymphocyte Stimulator polypeptides of the invention, their preparation,and compositions (preferably, pharmaceutical compositions) containingthem. In specific embodiments, the antibodies of the invention bind BLymphocyte Stimulator monomers, dimers, trimers or tetramers. Inadditional embodiments, the antibodies of the invention bind at leastdimers, at least trimers, or at least tetramers of B LymphocyteStimulator.

Multimeric B Lymphocyte Stimulator bound by the antibodies of theinvention may be homomers or heteromers. A B Lymphocyte Stimulatorhomomer, refers to a multimer containing only B Lymphocyte Stimulatorpolypeptides (including B Lymphocyte Stimulator fragments, variants, andfusion proteins, as described herein). These homomers may contain BLymphocyte Stimulator polypeptides having identical or different aminoacid sequences. In specific embodiments, the antibodies of the inventionbind a B Lymphocyte Stimulator homodimer (e.g., containing two BLymphocyte Stimulator polypeptides having identical or different aminoacid sequences) or a B Lymphocyte Stimulator homotrimer (e.g.,containing three B Lymphocyte Stimulator polypeptides having identicalor different amino acid sequences). In a preferred embodiment, theantibodies of the invention bind homotrimers of B Lymphocyte Stimulator.In additional embodiments, the antibodies of the invention bind ahomomeric B Lymphocyte Stimulator multimer which is at least ahomodimer, at least a homotrimer, or at least a homotetramer.

Heteromeric B Lymphocyte Stimulator refers to a multimer containingheterologous polypeptides (i.e., polypeptides of a different protein) inaddition to the B Lymphocyte Stimulator polypeptides of the invention.In a specific embodiment, the antibodies of the invention bind a BLymphocyte Stimulator heterodimer, a heterotrimer, or a heterotetramer.In additional embodiments, the antibodies of the invention bind aheteromeric B Lymphocyte Stimulator multimer which is at least aheterodimer, at least a heterotrimer, or at least a heterotetramer. Inhighly preferred embodiments, the antibodies of the invention bind aheterotrimer comprising both B Lymphocyte Stimulator polypeptides andAPRIL polypeptides (SEQ ID NO:3239; GenBank Accession No. AF046888; PCTInternational Publication Number WO97/33902; J. Exp. Med.188(6):1185-1190) or fragments or variants thereof. In other highlypreferred embodiments, the antibodies of the invention bind aheterotrimer comprising one B Lymphocyte Stimulator polypeptide(including fragments or variants) and two APRIL polypeptides (includingfragments or variants). In still other highly preferred embodiments, theantibodies of the invention bind a heterotrimer comprising two BLymphocyte Stimulator polypeptides (including fragments or variants) andone APRIL polypeptide (including fragments or variants). In a furthernonexclusive embodiment, the heteromers bound by the antibodies of theinvention contain CD40 ligand polypeptide sequence(s), or biologicallyactive fragment(s) or variant(s) thereof.

In particularly preferred embodiments, the antibodies of the inventionbind homomeric, especially homotrimeric, B Lymphocyte Stimulatorpolypeptides, wherein the individual protein components of the multimersconsist of the mature form of B Lymphocyte Stimulator (e.g., amino acidsresidues 134-285 of SEQ ID NO:3228, or amino acids residues 134-266 ofSEQ ID NO:3229) or fragments or variants thereof. In other specificembodiments, antibodies of the invention bind heteromeric, especiallyheterotrimeric, B Lymphocyte Stimulator polypeptides such as aheterotrimer containing two B Lymphocyte Stimulator polypeptides and oneAPRIL polypeptide or a heterotrimer containing one B LymphocyteStimulator polypeptide and two APRIL polypeptides, and wherein theindividual protein components of the B Lymphocyte Stimulator heteromerconsist of the mature extracellular soluble portion of either BLymphocyte Stimulator (e.g., amino acids residues 134-285 of SEQ IDNO:3228, or amino acids residues 134-266 of SEQ ID NO:3229) or fragmentsor variants thereof, or the mature extracellular soluble portion APRIL(e.g., amino acid residues 105-250 of SEQ ID NO:3239) or fragments orvariants thereof.

In specific embodiments, the antibodies of the invention bindconformational epitopes of a B Lymphocyte Stimulator monomeric protein.In specific embodiments, the antibodies of the invention bindconformational epitopes of a B Lymphocyte Stimulator multimeric,especially trimeric, protein. In other embodiments, antibodies of theinvention bind conformational epitopes that arise from the juxtapositionof B Lymphocyte Stimulator with a heterologous polypeptide, such asmight be present when B Lymphocyte Stimulator forms heterotrimers (e.g.,with APRIL polypeptides (e.g., SEQ ID SEQ ID NO:3239)), or in fusionproteins between B Lymphocyte Stimulator and a heterologous polypeptide.

In a specific embodiment, antibodies of the invention that specificallybind heterotrimers containing at least one B Lymphocyte Stimulatorpolypeptide and at least one APRIL polypeptide, comprise all or aportion of SEQ ID NOS: 1881 or 1884 (e.g., one or more CDR regions, a VHdomain or a VL domain). In a specific embodiment, the heterotrimerscontaining at least one B Lymphocyte Stimulator polypeptide and at leastone APRIL polypeptide comprise two B Lymphocyte Stimulator polypeptidesand one APRIL polypeptide. In a specific embodiment, the heterotrimerscontaining at least one B Lymphocyte Stimulator polypeptide and at leastone APRIL polypeptide comprise one B Lymphocyte Stimulator polypeptideand two APRIL polypeptides.

In a specific embodiment, antibodies of the invention that specificallybind heterotrimers containing at least one B Lymphocyte Stimulatorpolypeptide and at least one APRIL polypeptide, comprise all or aportion of any one of SEQ ID NOS: 3240-3247 (e.g., one or more CDRregions, a VH domain or a VL domain). The sequences of SEQ ID NOS:3240-3247 are presented after Table 1 just prior to the claims. In aspecific embodiment, the heterotrimers containing at least one BLymphocyte Stimulator polypeptide and at least one APRIL polypeptidecomprise two B Lymphocyte Stimulator polypeptides and one APRILpolypeptide. In a specific embodiment, the heterotrimers containing atleast one B Lymphocyte Stimulator polypeptide and at least one APRILpolypeptide comprise one B Lymphocyte Stimulator polypeptide and twoAPRIL polypeptides.

B Lymphocyte Stimulator multimers bound by the antibodies of theinvention may be the result of hydrophobic, hydrophilic, ionic and/orcovalent associations and/or may be indirectly linked, by for example,liposome formation. Thus, in one embodiment, B Lymphocyte Stimulatormultimers, such as, for example, homodimers or homotrimers, are formedwhen polypeptides of the invention contact one another in solution. Inanother embodiment, B Lymphocyte Stimulator heteromultimers, such as,for example, B Lymphocyte Stimulator heterotrimers or B LymphocyteStimulator heterotetramers, are formed when polypeptides of theinvention contact antibodies to the polypeptides of the invention(including antibodies to the heterologous polypeptide sequence in afusion protein of the invention) in solution. In other embodiments, BLymphocyte Stimulator multimers are formed by covalent associations withand/or between the B Lymphocyte Stimulator polypeptides of theinvention. Such covalent associations may involve one or more amino acidresidues contained in the polypeptide sequence (e.g., that recited inSEQ ID NO:3228 or SEQ ID NO:3229). In one instance, the covalentassociations are cross-linking between cysteine residues located withinthe polypeptide sequences which interact in the native (i.e., naturallyoccurring) polypeptide. In another instance, the covalent associationsare the consequence of chemical or recombinant manipulation.Alternatively, such covalent associations may involve one or more aminoacid residues contained in the heterologous polypeptide sequence in a BLymphocyte Stimulator fusion protein. In one example, covalentassociations are between the heterologous sequence contained in a fusionprotein (see, e.g., U.S. Pat. No. 5,478,925). In a specific example, thecovalent associations are between the heterologous sequence contained ina B Lymphocyte Stimulator-Fc fusion protein. In another specificexample, covalent associations of fusion proteins of the invention arebetween heterologous polypeptide sequence from another TNF familyligand/receptor member that is capable of forming covalently associatedmultimers, such as for example, oseteoprotegerin (see, e.g.,International Publication No. WO 98/49305, the contents of which areherein incorporated by reference in its entirety). In another specificexample, covalent associations of fusion proteins of the invention arebetween heterologous polypeptide sequence from CD40L, or a solublefragment thereof. In another embodiment, two or B Lymphocyte Stimulatorpolypeptides are joined through synthetic linkers (e.g., peptide,carbohydrate or soluble polymer linkers). Examples include those peptidelinkers described in U.S. Pat. No. 5,073,627 (hereby incorporated byreference). Proteins comprising multiple B Lymphocyte Stimulatorpolypeptides separated by peptide linkers may be produced usingconventional recombinant DNA technology.

In one embodiment, antibodies of the invention immunospecifically bind aB Lymphocyte Stimulator polypeptide having the amino acid sequence ofSEQ ID NO:3228 or as encoded by the cDNA clone contained in ATCC™ No.97768, or a polypeptide comprising a portion (i.e., a fragment) of theabove polypeptides. In another embodiment, the invention provides anantibody that binds an isolated B Lymphocyte Stimulator polypeptidehaving the amino acid sequence of SEQ ID NO:3229 or the amino acidsequence encoded by the cDNA clone contained in ATCC™ No. 203518, or anantibody that binds polypeptide comprising a portion (i.e, fragment) ofthe above polypeptides.

Antibodies of the invention that bind B Lymphocyte Stimulatorpolypeptides may bind them in as isolated polypeptides, in theirnaturally occurring state and/or their native conformation. By “isolatedpolypeptide” is intended a polypeptide removed from its nativeenvironment. Thus, a polypeptide produced by and/or contained within arecombinant host cell is considered isolated for purposes of the presentinvention. Also intended as an “isolated polypeptide” are polypeptidesthat have been purified, partially or substantially, from a recombinanthost cell. Thus, antibodies of the present invention may bindrecombinantly produced B Lymphocyte Stimulator polypeptides.

Antibodies of the present invention may also bind B LymphocyteStimulator expressed on the surface of a cell, wherein said B LymphocyteStimulator polypeptide is encoded by a polynucleotide encoding aminoacids 1 to 285 of SEQ ID NO:2 operably associated with a regulatorysequence that controls expression of said polynucleotide. In certainembodiments, said B Lymphocyte Stimulator polypeptide expressed on thesurface of a cell is a recombinant B Lymphocyte Stimulator polypeptide.In other embodiments, said B Lymphocyte Stimulator polypeptide expressedon the surface of the cell is a naturally occurring B LymphocyteStimulator polypeptide. As a non-limiting example, an antibody of theinvention may bind a B Lymphocyte Stimulator expressed on the surface ofthe cell wherein Lys-132 and/or Arg-133 of the B Lymphocyte Stimulatorsequence shown in SEQ ID NO:3228 is mutated to another amino acidresidue, or deleted altogether, thereby preventing or diminishingrelease of the soluble form of B Lymphocyte Stimulator from cellsexpressing B Lymphocyte Stimulator.

Antibodies of the present invention may also bind B LymphocyteStimulator secreted by a cell, wherein said B Lymphocyte Stimulatorpolypeptide is encoded by a polynucleotide encoding amino acids 1 to 285of SEQ ID NO:2 operably associated with a regulatory sequence thatcontrols expression of said polynucleotide. In certain embodiments, saidB Lymphocyte Stimulator polypeptide secreted by a cell is a recombinantB Lymphocyte Stimulator polypeptide. In other embodiments, said BLymphocyte Stimulator polypeptide secreted by a cell is a naturallyoccurring B Lymphocyte Stimulator polypeptide.

Antibodies of the present invention immunospecifically bind topolypeptides comprising or alternatively, consisting of, the amino acidsequence of SEQ ID NO:3228, encoded by the cDNA contained in the plasmidhaving ATCC™ accession number 97768, or encoded by nucleic acids whichhybridize (e.g., under stringent hybridization conditions) to thenucleotide sequence contained in the deposited clone. Antibodies of thepresent invention also bind to fragments of the amino acid sequence ofSEQ ID NO:3228, encoded by the cDNA contained in the plasmid havingATCC™ accession number 97768, or encoded by nucleic acids whichhybridize (e.g., under stringent hybridization conditions) to thenucleotide sequence contained in the deposited clone.

Additionally, antibodies of the present invention bind polypeptidescomprising or alternatively, consisting of, the amino acid sequence ofSEQ ID NO:3229, encoded by the cDNA contained in the plasmid havingATCC™ accession number 203518, or encoded by nucleic acids whichhybridize (e.g., under stringent hybridization conditions) to thenucleotide sequence contained in the deposited clone. Antibodies of thepresent invention also bind to fragments of the amino acid sequence ofSEQ ID NO:3229, encoded by the cDNA contained in the plasmid havingATCC™ accession number 203518, or encoded by nucleic acids whichhybridize (e.g., under stringent hybridization conditions) to thenucleotide sequence contained in the deposited clone.

In addition, antibodies of the invention bind polypeptides orpolypeptide fragments comprising or alternatively, consisting of, anamino acid sequence contained in SEQ ID NOS: 3230 through 3237.

In specific embodiments, the antibodies of the present inventionimmunospecifically bind polypeptide fragments including polypeptidescomprising or alternatively, consisting of, an amino acid sequencecontained in SEQ ID NO:3228, encoded by the cDNA contained in thedeposited clone, or encoded by nucleic acids which hybridize (e.g.,under stringent hybridization conditions) to the nucleotide sequencecontained in the deposited clone. Protein fragments may be“free-standing,” or comprised within a larger polypeptide of which thefragment forms a part or region, most preferably as a single continuousregion. Representative examples of polypeptide fragments that may bebound by the antibodies of the present invention, include, for example,fragments that comprise or alternatively, consist of from about aminoacid residues: 1 to 50, 51 to 100, 101 to 150, 151 to 200, 201 to 250,and/or 251 to 285 of SEQ ID NO:3228. Moreover, polypeptide fragments canbe at least 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140,150, 175 or 200 amino acids in length.

In specific embodiments, antibodies of the present invention bindpolypeptide fragments comprising, or alternatively consisting of, aminoacid residues: 1-46, 31-44, 47-72, 73-285, 73-83, 94-102, 148-152,166-181, 185-209, 210-221, 226-237, 244-249, 253-265, and/or 277-285 ofSEQ ID NO:3228. In a specific embodiment, antibodies of the inventionbind an epitope comprising amino acids 165-171 of SEQ ID NO:3228. Inanother embodiment, the CDRs of antibodies of the invention makecontacts with one or more amino acids in the sequence of amino acids165-171 of SEQ ID NO:3228. In another embodiment, antibodies of theinvention whose CDRs make contact with one or more amino acids in thesequence of amino acids 165-171 of SEQ ID NO:3228 disrupt B LymphocyteStimulator-B Lymphocyte Stimulator receptor interactions.

It will be recognized by one of ordinary skill in the art that mutationstargeted to regions of a B Lymphocyte Stimulator polypeptide of SEQ IDNO:3228 which encompass the nineteen amino acid residue insertion whichis not found in the B Lymphocyte Stimulator polypeptide sequence of SEQID NO:3229 (i.e., amino acid residues Val-142 through Lys-160 of thesequence of SEQ ID NO:3229) may affect the observed biologicalactivities of the B Lymphocyte Stimulator polypeptide. Morespecifically, a partial, non-limiting and non-exclusive list of suchresidues of the B Lymphocyte Stimulator polypeptide sequence which maybe targeted for mutation includes the following amino acid residues ofthe B Lymphocyte Stimulator polypeptide sequence as shown in SEQ IDNO:3228: V-142; T-143; Q-144; D-145; C-146; L-147; Q-148; L-149; I-150;A-151; D-152; S-153; E-154; T-155; P-156; T-157; I-158; Q-159; andK-160. Thus, in specific embodiments, antibodies of the presentinvention that bind B Lymphocyte Stimulator polypeptides which have oneor more mutations in the region from V-142 through K-160 of SEQ IDNO:3228 are contemplated.

Polypeptide fragments may be “free-standing,” or comprised within alarger polypeptide of which the fragment forms a part or region, mostpreferably as a single continuous region. Representative examples ofpolypeptide fragments that may be bound by antibodies of the presentinvention, include, for example, fragments that comprise oralternatively, consist of from about amino acid residues: 1 to 15,16-30, 31-46, 47-55, 56-72, 73-104, 105-163, 163-188, 186-210 and210-284 of the amino acid sequence disclosed in SEQ ID NO:3228.Additional representative examples of polypeptide fragments that may bebound by antibodies of the present invention, include, for example,fragments that comprise or alternatively, consist of from about aminoacid residues: 1 to 143, 1-150, 47-143, 47-150, 73-143, 73-150, 100-150,140-145, 142-148, 140-150, 140-200, 140-225, and 140-266 of the aminoacid sequence disclosed in SEQ ID NO:3229. Moreover, polypeptidefragments that may be bound by antibodies of the present invention, canbe at least 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140,150, 175 or 200 amino acids in length. In this context, “about” meansthe particularly recited ranges and ranges larger or smaller by several,a few, 5, 4, 3, 2 or 1 amino acid residues at either or both the amino-and carboxy-termini.

Additional preferred embodiments encompass antibodies that bindpolypeptide fragments comprising, or alternatively consisting of, thepredicted intracellular domain of B Lymphocyte Stimulator (e.g., aminoacid residues 1-46 of SEQ ID NO:3228), the predicted transmembranedomain of B Lymphocyte Stimulator (e.g., amino acid residues 47-72 ofSEQ ID NO:3228), the predicted extracellular domain of B LymphocyteStimulator (e g , amino acid residues 73-285 of SEQ ID NO:3228), themature soluble extracellular domain of B Lymphocyte Stimulator (e.g.,amino acids residues 134-285 of SEQ ID NO:3228), the predicted TNFconserved domain of B Lymphocyte Stimulator (e.g., amino acids 191 to284 of SEQ ID NO:3228), and a polypeptide comprising, or alternatively,consisting of the predicted intracellular domain fused to the predictedextracellular domain of B Lymphocyte Stimulator (amino acid residues1-46 fused to amino acid residues 73-285 of SEQ ID NO:3228).

Further additional preferred embodiments encompass polypeptide fragmentscomprising, or alternatively consisting of, the predicted intracellulardomain of B Lymphocyte Stimulator (amino acid residues 1-46 of SEQ IDNO:3229), the predicted transmembrane domain of B Lymphocyte Stimulator(amino acid residues 47-72 of SEQ ID NO:3229), the predictedextracellular domain of B Lymphocyte Stimulator (amino acid residues73-266 of SEQ ID NO:3229), the predicted TNF conserved domain of BLymphocyte Stimulator (amino acids 172 to 265 of SEQ ID NO:3229), and apolypeptide comprising, or alternatively, consisting of the predictedintracellular domain fused to the predicted extracellular domain of BLymphocyte Stimulator (amino acid residues 1-46 fused to amino acidresidues 73-266 of SEQ ID NO:3229).

Certain additional embodiments of the invention encompass antibodiesthat bind polypeptide fragments comprising, or alternatively consistingof, the predicted beta-pleated sheet regions of the B LymphocyteStimulator polypeptides of SEQ ID NO:3228 and SEQ ID NO:3229. Thesepolypeptide fragments comprising the beta-pleated sheets of B LymphocyteStimulator comprise, or alternatively consist of, amino acid residuesGln-144 to Ala-151, Phe-172 to Lys-173, Ala-177 to Glu-179, Asn-183 toIle-185, Gly-191 to Lys-204, His-210 to Val-219, Leu-226 to Pro-237,Asn-242 to Ala-251, Gly-256 to Ile-263 and/or Val-276 to Leu-284 of SEQID NO:3228. In another, nonexclusive embodiment, these polypeptidefragments comprising the beta-pleated sheets of B Lymphocyte Stimulatorcomprise, or alternatively consist of, amino acid residues Phe-153 toLys-154, Ala-158 to Glu-160, Asn-164 to Ile-166, Gly-172 to Lys-185,His-191 to Val-200, Leu-207 to Pro-218, Asn-223 to Ala-232, Gly-237 toIle-244 and/or Val-257 to Leu-265 of SEQ ID NO:3229.

A partial, non-limiting, and exemplary list of polypeptides that may bebound by the antibodies of the invention includes polypeptides thatcomprise, or alternatively consist of, combinations of amino acidsequences of the invention includes, for example, [Met-1 to Lys-113]fused to [Leu-114 to Thr-141] fused to [Val-142 to Lys-160] fused to[Gly-161 to Gln-198] fused to [Val-199 to Ala-248] fused to [Gly-249 toLeu-285] of SEQ ID NO:3228; or [Met-1 to Lys-113] fused to [Val-142 toLys-160] fused to [Gly-161 to Gln-198] fused to [Val-199 to Ala-248]fused to [Gly-249 to Leu-285] of SEQ ID NO:3228; or [Met-1 to Lys-113]fused to [Leu-114 to Thr-141] fused to [Val-142 to Lys-160] fused to[Gly-161 to Gln-198] fused to [Gly-249 to Leu-285] of SEQ ID NO:3228.Other combinations of amino acids sequences that may be bound by theantibodies of the invention may include the polypeptide fragments in anorder other than that recited above (e.g., [Leu-114 to Thr-141] fused to[Val-199 to Ala-248] fused to [Gly-249 to Leu-285] fused to [Val-142 toLys-160] of (SEQ ID NO:3228). Other combinations of amino acidssequences that may be bound by the antibodies of the invention may alsoinclude heterologous polypeptide fragments as described herein and/orother polypeptides or polypeptide fragments of the present invention(e.g., [Met-1 to Lys-113] fused to [Leu-114 to Thr-141] fused to[Val-142 to Lys-160] fused to [Gly-161 to Gln-198] fused to [Gly-249 toLeu-285] of SEQ ID NO:3228 fused to a FLAG tag; or [Met-1 to Lys-113] ofSEQ ID NO:3228 fused to [Leu-114 to Thr-141] of SEQ ID NO:3228 fused to[Glu-135 to Asn-165] of SEQ ID NO:39 fused to [Val-142 to Lys-160] ofSEQ ID NO:3228 fused to [Gly-161 to Gln-198] of SEQ ID NO:3228 fused to[Val-199 to Ala-248] of SEQ ID NO:3228 fused to [Gly-249 to Leu-285] ofSEQ ID NO:3228).

A partial, non-limiting, and exemplary list of polypeptides that may bebound by the antibodies of the invention includes polypeptides thatcomprise, or alternatively consist of, combinations of amino acidsequences includes, for example, [Met-1 to Lys-113] fused to [Leu-114 toThr-141] fused to [Gly-142 to Gln-179] fused to [Val-180 to Ala-229]fused to [Gly-230 to Leu-266] of SEQ ID NO:3229; [Met-1 to Lys-113]fused to [Gly-142 to Gln-179] fused to [Val-180 to Ala-229] fused to[Gly-230 to Leu-266] of SEQ ID NO:3229; or [Met-1 to Lys-113] fused to[Leu-114 to Thr-141] fused to [Gly-142 to Gln-179] fused to [Gly-230 toLeu-266] of SEQ ID NO:3229. Other of amino acids sequences that may bebound by the antibodies of the invention combinations may include thepolypeptide fragments in an order other than that recited above (e.g.,[Leu-114 to Thr-141] fused to [Val-180 to Ala-229] fused to [Gly-230 toLeu-266] fused to [Gly-142 to Gln-179] of SEQ ID NO:3229). Othercombinations of amino acid sequences that may be bound by the antibodiesof the invention may also include heterologous polypeptide fragments asdescribed herein and/or other polypeptides or polypeptide fragments ofthe present invention (e.g., [Met-1 to Lys-113] fused to [Leu-114 toThr-141] fused to [Gly-142 to Gln-179] fused to [Gly-230 to Leu-266] ofSEQ ID NO:3229 fused to a FLAG tag (SEQ ID NO:3238) or, [Met-1 toLys-113] of SEQ ID NO:3229 fused to [Leu-114 to Thr-141] of SEQ IDNO:3229 fused to [Glu-135 to Asn-165] of SEQ ID NO:39 fused to [Gly-142to Gln-179] of SEQ ID NO:3229 fused to [Val-180 to Ala-229] of SEQ IDNO:3229 fused to [Gly-230 to Leu-266] of SEQ ID NO:3229.

Additional embodiments of the invention encompass antibodies that bind BLymphocyte Stimulator polypeptide fragments comprising, or alternativelyconsisting of, functional regions of polypeptides of the invention, suchas the Garnier-Robson alpha-regions, beta-regions, turn-regions, andcoil-regions, Chou-Fasman alpha-regions, beta-regions, and coil-regions,Kyte-Doolittle hydrophilic regions and hydrophobic regions, Eisenbergalpha- and beta-amphipathic regions, Karplus-Schulz flexible regions,Emini surface-forming regions and Jameson-Wolf regions of high antigenicindex set out in Tables 9 and 10 and as described herein. In a preferredembodiment, the polypeptide fragments bound by the antibodies of theinvention are antigenic (i.e., containing four or more contiguous aminoacids having an antigenic index of greater than or equal to 1.5, asidentified using the default parameters of the Jameson-Wolf program) ofa complete (i.e., full-length) B Lymphocyte Stimulator polypeptide(e.g., SEQ ID NOS:3228 and 3229).

The data representing the structural or functional attributes of the BLymphocyte Stimulator polypeptide of SEQ ID NO:3228 (Table 9) or the BLymphocyte Stimulator polypeptide of SEQ ID NO:3229 (Table 10), asdescribed above, was generated using the various modules and algorithmsof the DNA*STAR set on default parameters. Column I represents theresults of a Gamier-Robson analysis of alpha helical regions; Column IIrepresents the results of a Chou-Fasman analysis of alpha helicalregions; Column III represents the results of a Gamier Robson analysisof beta sheet regions; Column IV represents the results of a Chou-Fasmananalysis of beta sheet regions; Column V represents the results of aGamier Robson analysis of turn regions; Column VI represents the resultsof a Chou-Fasman analysis of turn regions; Column VII represents theresults of a Gamier Robson analysis of coil regions; Column VIIIrepresents a Kyte-Doolittle hydrophilicity plot; Column IX represents aHopp-Woods hydrophobicity plot; Column X represents the results of anEisenberg analysis of alpha amphipathic regions; Column XI representsthe results of an Eisenberg analysis of beta amphipathic regions; ColumnXII represents the results of a Karplus-Schultz analysis of flexibleregions; Column XIII represents the Jameson-Wolf antigenic index score;and Column XIV represents the Emini surface probability plot.

In a preferred embodiment, the data presented in columns VIII, IX, XIII,and XIV of Tables 9 and 10 can be used to determine regions of the BLymphocyte Stimulator polypeptide of SEQ ID NO:3228 (Table 9) or the BLymphocyte Stimulator polypeptide of SEQ ID NO:3229 (Table 10) whichexhibit a high degree of potential for antigenicity. Regions of highantigenicity are determined from the data presented in columns VIII, IX,XIII, and/or XIV by choosing values which represent regions of thepolypeptide which are likely to be exposed on the surface of thepolypeptide in an environment in which antigen recognition may occur inthe process of initiation of an immune response.

The above-mentioned preferred regions set out in Tables 9 and 10include, but are not limited to, regions of the aforementioned typesidentified by analysis of the amino acid sequence set out in SEQ IDNO:2. As set out in Tables 9 and 10, such preferred regions includeGarnier-Robson alpha-regions, beta-regions, turn-regions, andcoil-regions, Chou-Fasman alpha-regions, beta-regions, and turn-regions,Kyte-Doolittle hydrophilic regions, Eisenberg alpha- andbeta-amphipathic regions, Karplus-Schulz flexible regions, Jameson-Wolfregions of high antigenic index and Emini surface-forming regions.Preferably, antibodies of the present invention bind B LymphocyteStimulator polypeptides or B Lymphocyte Stimulator polypeptide fragmentsand variants comprising regions of B Lymphocyte Stimulator that combineseveral structural features, such as several (e.g., 1, 2, 3, or 4) ofthe same or different region features set out above and in Tables 9 and10.

TABLE 9 Res Position I II III IV V VI VII VIII IX X XI XII XIII XIV Met1 A . . . . . . 0.73 −0.71 . . . 0.95 1.39 Asp 2 A . . . . T . 1.12−0.66 * . . 1.15 1.56 Asp 3 A . . . . T . 1.62 −1.09 * . . 1.15 2.12 Ser4 A . . . . T . 2.01 −1.51 . . . 1.15 4.19 Thr 5 A . . . . T . 2.40−2.13 . . F 1.30 4.35 Glu 6 A A . . . . . 2.70 −1.73 * * F 0.90 4.51 Arg7 A A . . . . . 2.81 −1.34 * * F 0.90 4.51 Glu 8 A A . . . . . 2.00−1.73 * * F 0.90 6.12 Gln 9 A A . . . . . 1.99 −1.53 * * F 0.90 2.91 Ser10 A . . B . . . 2.00 −1.04 * * F 0.90 2.15 Arg 11 A . . B . . . 1.33−0.66 * * F 0.90 1.66 Leu 12 A . . B . . . 0.41 −0.09 * * F 0.45 0.51Thr 13 A . . B . . . 0.46 0.20 * * F −0.15 0.32 Ser 14 A A . . . . .0.50 −0.19 * * . 0.30 0.32 Cys 15 A A . . . . . 0.91 −0.19 * * . 0.300.78 Leu 16 A A . . . . . 0.80 −0.87 * * F 0.90 1.06 Lys 17 A A . . . .. 1.61 −1.36 . * F 0.90 1.37 Lys 18 A A . . . . . 1.32 −1.74 . * F 0.904.44 Arg 19 A A . . . . . 1.67 −1.70 . * F 0.90 5.33 Glu 20 A A . . . .. 1.52 −2.39 . * F 0.90 5.33 Glu 21 A A . . . . . 2.38 −1.70 . * F 0.902.20 Met 22 A A . . . . . 2.33 −1.70 . * F 0.90 2.24 Lys 23 A A . . . .. 1.62 −1.70 * * F 0.90 2.24 Leu 24 A A . . . . . 0.66 −1.13 * * F 0.750.69 Lys 25 A A . . . . . 0.36 −0.49 . * F 0.45 0.52 Glu 26 A A . B . .. −0.53 −0.71 * * . 0.60 0.35 Cys 27 A A . B . . . −0.74 −0.03 * * .0.30 0.30 Val 28 A A . B . . . −1.00 −0.03 * * . 0.30 0.12 Ser 29 A A .B . . . −0.08 0.40 * * . −0.30 0.11 Ile 30 A . . B . . . −0.08 0.40 * *. −0.30 0.40 Leu 31 A . . B . . . −0.08 −0.17 * . . 0.45 1.08 Pro 32 . .. B . . C 0.29 −0.81 * . F 1.10 1.39 Arg 33 . . . . T . . 0.93 −0.81 . *F 1.50 2.66 Lys 34 . . . . T . . 0.93 −1.07 . . F 1.84 4.98 Glu 35 . . .. . . C 0.97 −1.37 * * F 1.98 4.32 Ser 36 . . . . . T C 1.89 −1.16 * * F2.52 1.64 Pro 37 . . . . . T C 1.80 −1.16 * * F 2.86 1.60 Ser 38 . . . .T T . 1.39 −0.77 * . F 3.40 1.24 Val 39 A . . . . T . 1.39 −0.39 . * F2.36 1.24 Arg 40 A . . . . . . 1.39 −0.77 * * F 2.46 1.60 Ser 41 A . . .. . . 1.34 −1.20 * * F 2.46 2.00 Ser 42 . . . . T T . 1.60 −1.16 . * F3.06 2.67 Lys 43 . . . . T T . 1.09 −1.80 . * F 3.06 2.72 Asp 44 . . . .T T . 1.13 −1.11 * * F 3.40 1.67 Gly 45 A . . . . T . 0.43 −0.81 * * F2.66 1.03 Lys 46 A A . . . . . 0.14 −0.70 . . F 1.77 0.52 Leu 47 A A . .. . . 0.13 −0.20 * . . 0.98 0.31 Leu 48 A A . . . . . −0.72 0.29 * . .0.04 0.46 Ala 49 A A . . . . . −1.53 0.54 . * . −0.60 0.19 Ala 50 A A .. . . . −2.00 1.23 . . . −0.60 0.19 Thr 51 A A . . . . . −2.63 1.23 . .. −0.60 0.19 Leu 52 A A . . . . . −2.63 1.04 . . . −0.60 0.19 Leu 53 A A. . . . . −2.63 1.23 . . . −0.60 0.15 Leu 54 A A . . . . . −2.34 1.41 .. . −0.60 0.09 Ala 55 A A . . . . . −2.42 1.31 . . . −0.60 0.14 Leu 56 AA . . . . . −2.78 1.20 . . . −0.60 0.09 Leu 57 A . . . . T . −2.78 1.09. . . −0.20 0.06 Ser 58 A . . . . T . −2.28 1.09 . . . −0.20 0.05 Cys 59A . . . . T . −2.32 1.07 . . . −0.20 0.09 Cys 60 A . . . . T . −2.591.03 . . . −0.20 0.08 Leu 61 . . B B . . . −2.08 0.99 . . . −0.60 0.04Thr 62 . . B B . . . −1.97 0.99 . . . −0.60 0.11 Val 63 . . B B . . .−1.91 1.20 . . . −0.60 0.17 Val 64 . . B B . . . −1.24 1.39 . . . −0.600.33 Ser 65 . . B B . . . −1.43 1.10 . . . −0.60 0.40 Phe 66 A . . B . .. −1.21 1.26 . . . −0.60 0.40 Tyr 67 A . . B . . . −1.49 1.11 . . .−0.60 0.54 Gln 68 A . . B . . . −1.44 0.97 . . . −0.60 0.41 Val 69 A . .B . . . −0.59 1.27 . . . −0.60 0.39 Ala 70 A . . B . . . −0.63 0.89 . .. −0.60 0.43 Ala 71 A . . B . . . 0.07 0.56 . * . −0.60 0.25 Leu 72 A .. . . T . −0.50 0.16 . * . 0.10 0.55 Gln 73 A . . . . T . −1.09 0.20 . .F 0.25 0.45 Gly 74 A . . . . T . −0.53 0.20 . . F 0.25 0.45 Asp 75 A . .. . T . −0.76 0.09 . * F 0.25 0.73 Leu 76 A A . . . . . −0.06 0.09 . * F−0.15 0.35 Ala 77 A A . . . . . 0.17 −0.31 . * . 0.30 0.69 Ser 78 A A .. . . . 0.17 −0.24 . * . 0.30 0.42 Leu 79 A A . . . . . −0.30 −0.24 . *. 0.30 0.88 Arg 80 A A . . . . . −0.30 −0.24 . * . 0.30 0.72 Ala 81 A A. . . . . 0.17 −0.34 . * . 0.30 0.93 Glu 82 A A . . . . . 0.72 −0.30 . *. 0.45 1.11 Leu 83 A A . . . . . 0.99 −0.49 . * . 0.30 0.77 Gln 84 A A .. . . . 1.21 0.01 . * . −0.15 1.04 Gly 85 A A . . . . . 1.10 0.01 * * .−0.30 0.61 His 86 A A . . . . . 1.73 0.01 * * . −0.15 1.27 His 87 A A .. . . . 0.92 −0.67 . * . 0.75 1.47 Ala 88 A A . . . . . 1.52 −0.39 . * .0.45 1.22 Glu 89 A A . . . . . 0.93 −0.39 . . . 0.45 1.39 Lys 90 A A . .. . . 0.93 −0.39 * . F 0.60 1.03 Leu 91 A . . . . T . 0.38 −0.46 * . .0.85 1.01 Pro 92 A . . . . T . 0.07 −0.46 . . . 0.70 0.59 Ala 93 A . . .. T . 0.07 −0.03 . . . 0.70 0.29 Gly 94 A . . . . T . −0.14 0.47 . . .−0.20 0.36 Ala 95 A . . . . . . −0.14 0.21 . * . −0.10 0.36 Gly 96 A . .. . . . 0.08 −0.21 . . F 0.65 0.71 Ala 97 A . . . . . . −0.06 −0.21 . .F 0.65 0.72 Pro 98 A . . . . . . −0.28 −0.21 . * F 0.65 0.71 Lys 99 A A. . . . . 0.07 −0.03 . . F 0.45 0.59 Ala 100 A A . . . . . 0.66 −0.46 .. F 0.60 1.01 Gly 101 A A . . . . . 0.41 −0.96 . . F 0.90 1.13 Leu 102 AA . . . . . 0.79 −0.89 . . F 0.75 0.57 Glu 103 A A . . . . . 0.41−0.46 * . F 0.45 0.88 Glu 104 A A . . . . . −0.49 −0.46 * . F 0.45 0.89Ala 105 A A . . . . . −0.21 −0.24 . . . 0.30 0.81 Pro 106 A A . . . . .−0.46 −0.44 . . . 0.30 0.67 Ala 107 A A . . . . . 0.01 0.06 . . . −0.300.39 Val 108 A A . . . . . −0.80 0.49 . * . −0.60 0.38 Thr 109 A A . . .. . −0.76 0.67 . * . −0.60 0.20 Ala 110 A A . . . . . −1.06 0.24 * * .−0.30 0.40 Gly 111 A A . . . . . −1.54 0.43 * * . −0.60 0.38 Leu 112 A A. . . . . −0.96 0.57 * * . −0.60 0.23 Lys 113 . A B . . . . −0.310.09 * * . −0.30 0.39 Ile 114 . A B . . . . −0.21 0.01 * . . −0.30 0.61Phe 115 . A B . . . . −0.21 0.01 * . . 0.15 1.15 Glu 116 . A . . . . C−0.08 −0.17 * . F 1.25 0.58 Pro 117 . A . . . . C 0.39 0.26 * * F 1.101.28 Pro 118 . . . . . . C 0.34 −0.00 . . F 2.20 1.47 Ala 119 . . . . .T C 0.89 −0.79 . * F 3.00 1.47 Pro 120 . . . . . T C 1.59 −0.36 . * F2.25 0.94 Gly 121 . . . . T T . 1.29 −0.39 . * F 2.15 0.98 Glu 122 . . .. T T . 1.20 −0.43 . . F 2.00 1.30 Gly 123 . . . . . . C 1.41 −0.54 . .F 1.60 1.12 Asn 124 . . . . . T C 2.00 −0.57 . . F 1.50 1.97 Ser 125 . .. . . T C 1.91 −0.60 . * F 1.50 1.82 Ser 126 . . . . . T C 2.37 −0.21. * F 1.54 2.47 Gln 127 . . . . . T C 2.37 −0.64 . * F 2.18 3.01 Asn 128. . . . . . C 2.76 −0.64 . . F 2.32 3.61 Ser 129 . . . . . T C 2.87−1.03 . . F 2.86 5.39 Arg 130 . . . . T T . 2.58 −1.41 * . F 3.40 6.09Asn 131 . . . . T T . 2.02 −1.31 * . F 3.06 3.83 Lys 132 . . . . T T .2.02 −1.07 * . F 2.72 2.12 Arg 133 . . . . T . . 1.68 −1.06 * . F 2.181.88 Ala 134 . . . . . . C 1.77 −0.63 * . F 1.64 1.15 Val 135 . . . . .. C 1.66 −0.60 * . F 1.49 0.89 Gln 136 . . . . . . C 1.66 −0.60 * . F1.83 0.79 Gly 137 . . . . . T C 1.30 −0.60 * . F 2.52 1.35 Pro 138 . . .. . T C 0.33 −0.61 * . F 2.86 2.63 Glu 139 . . . . T T . 0.61 −0.61 * .F 3.40 1.13 Glu 140 A . . . . T . 1.47 −0.53 * . F 2.66 1.64 Thr 141 A .. . . . . 1.47 −0.56 . . F 2.12 1.84 Val 142 A . . . . . . 1.14 −0.99 .. F 1.78 1.77 Thr 143 A . . . . T . 0.54 −0.41 . . F 1.19 0.55 Gln 144 A. . . . T . 0.54 0.27 * . F 0.25 0.31 Asp 145 A . . . . T . −0.27 0.19 *. F 0.25 0.73 Cys 146 A . . . . T . −0.84 0.23 * . . 0.10 0.42 Leu 147 AA . . . . . −0.58 0.43 * . . −0.60 0.17 Gln 148 A A . . . . . −0.270.53 * . . −0.60 0.10 Leu 149 A A . . . . . −0.57 0.53 * * . −0.30 0.32Ile 150 A A . . . . . −0.57 0.34 * . . 0.30 0.52 Ala 151 . A . . . . C−0.21 −0.34 . * . 1.40 0.52 Asp 152 . . . . T T . 0.39 −0.26 . * F 2.450.91 Ser 153 . . . . . T C 0.08 −0.51 . . F 3.00 2.00 Glu 154 . . . . .T C −0.00 −0.71 . . F 2.70 2.86 Thr 155 . . . . . T C 0.89 −0.53 * . F2.40 1.20 Pro 156 . . . B . . C 1.52 −0.13 * . F 1.56 1.55 Thr 157 . . .B T . . 1.18 −0.51 * . F 1.92 1.79 Ile 158 A . . B . . . 1.18 −0.09 . .F 1.08 1.23 Gln 159 . . . . T T . 0.93 −0.19 . . F 2.04 1.07 Lys 160 . .. . T T . 0.93 0.14 * . F 1.60 1.16 Gly 161 . . . . T T . 0.44 0.14 * .F 1.44 2.38 Ser 162 . . . . T T . −0.10 0.24 * . F 1.28 1.19 Tyr 163 . .. B T . . 0.58 0.49 * . . 0.12 0.44 Thr 164 . . B B . . . 0.29 0.91 * .. −0.44 0.69 Phe 165 . . B B . . . −0.57 1.40 * . . −0.60 0.54 Val 166 .. B B . . . −1.03 1.70 . . . −0.60 0.29 Pro 167 . . B B . . . −1.03 1.63. . . −0.60 0.16 Trp 168 A . . B . . . −1.49 1.53 . * . −0.60 0.25 Leu169 A . . B . . . −1.13 1.53 * . . −0.60 0.29 Leu 170 A . . B . . .−0.32 0.89 * . . −0.30 0.38 Ter 171 A . . . . . . 0.19 0.46 * . . 0.200.71 Phe 172 . . . . T . . 0.10 −0.03 * . . 1.80 0.85 Lys 173 . . . . TT . −0.20 −0.33 * . F 2.60 1.38 Arg 174 . . . . . T C −0.20 −0.51 . . F3.00 1.04 Gly 175 . . . . . T C 0.61 −0.21 . . F 2.25 0.99 Ser 176 A . .. . T . 0.91 −1.00 * . F 2.05 0.86 Ala 177 A A . . . . . 1.66 −1.00 * .F 1.35 0.76 Leu 178 A A . . . . . 1.61 −1.00 . . F 1.20 1.54 Glu 179 A A. . . . . 1.50 −1.43 . . F 0.90 1.98 Glu 180 A A . . . . . 1.89 −1.41 *. F 0.90 3.16 Lys 181 A A . . . . . 1.30 −1.91 * . F 0.90 7.66 Glu 182 AA . . . . . 1.08 −1.91 . . F 0.90 3.10 Asn 183 A A . . . . . 1.03−1.23 * * F 0.90 1.48 Lys 184 A A . . . . . 1.08 −0.59 * . F 0.75 0.55Ile 185 A A . . . . . 1.08 −0.59 * * . 0.60 0.63 Leu 186 A A . . . . .0.72 −0.59 * * . 0.60 0.68 Val 187 A A . . . . . 0.38 −0.50 . * . 0.300.49 Lys 188 A A . . . . . 0.13 −0.07 * * F 0.45 0.69 Glu 189 A . . . .T . −0.61 0.00 * * F 0.40 1.32 Thr 190 . . . . T T . −0.42 0.10 . * F0.80 1.54 Gly 191 . . . . T T . −0.50 0.24 * . F 0.65 0.67 Tyr 192 . . .. T T . 0.11 0.93 * * . 0.20 0.27 Phe 193 . . B B . . . −0.28 1.69 . . .−0.60 0.29 Phe 194 . . B B . . . −0.28 1.63 . * . −0.60 0.29 Ile 195 . .B B . . . −0.82 1.60 . . . −0.60 0.32 Tyr 196 . . B B . . . −1.29 1.49 .. . −0.60 0.28 Gly 197 . . . B T . . −1.29 1.39 . . . −0.20 0.26 Gln 198. . . B T . . −0.90 1.36 . . . −0.20 0.59 Val 199 . . . B . . C −0.201.16 . . . −0.40 0.54 Leu 200 . . . B . . C 0.73 0.40 . . . −0.10 0.92Tyr 201 . . . . T T . 0.67 −0.03 . . . 1.25 1.06 Thr 202 . . . . T T .0.77 0.06 . . F 0.80 2.06 Asp 203 . . . . T T . 0.18 0.17 . . F 0.803.91 Lys 204 A . . . . T . 0.43 −0.01 . . F 1.00 2.52 Thr 205 A A . . .. . 0.90 −0.16 . . F 0.60 1.73 Tyr 206 A A . . . . . 1.11 −0.21 . . .0.45 1.03 Ala 207 A A . . . . . 0.61 0.29 . . . −0.30 0.70 Met 208 A A .. . . . −0.28 0.97 . . . −0.60 0.40 Gly 209 A A . B . . . −0.32 1.17 * .. −0.60 0.18 His 210 A A . B . . . 0.10 0.81 * . . −0.60 0.31 Leu 211 AA . B . . . 0.39 0.31 . . . −0.30 0.61 Ile 212 A A . B . . . 1.02 −0.30. . . 0.45 1.22 Gln 213 A A . B . . . 0.77 −0.73 . * . 0.75 1.80 Arg 214A A . B . . . 1.08 −0.59 . * F 0.90 1.62 Lys 215 A A . B . . . 0.26−0.77 * * F 0.90 3.14 Lys 216 A A . B . . . 0.37 −0.81 . * F 0.90 1.35Val 217 . A B B . . . 0.91 −0.43 * * . 0.30 0.60 His 218 . A B B . . .0.91 −0.00 . * . 0.30 0.29 Val 219 . A B B . . . 0.80 −0.00 * * . 0.300.25 Phe 220 . . B B . . . −0.06 −0.00 * . . 0.30 0.57 Gly 221 A . . B .. . −0.40 0.04 . * . −0.30 0.35 Asp 222 A . . . . . . −0.36 −0.07 * . .0.50 0.63 Glu 223 A . . . . . . −1.18 −0.03 * . . 0.50 0.60 Leu 224 A .. B . . . −0.63 −0.17 . . . 0.30 0.45 Ser 225 A . . B . . . −0.74 −0.11. . . 0.30 0.39 Leu 226 A . . B . . . −1.10 0.57 . * . −0.60 0.18 Val227 A . . B . . . −0.99 1.36 . * . −0.60 0.19 Thr 228 A . . B . . .−1.66 0.67 * * . −0.60 0.28 Leu 229 A . . B . . . −1.73 0.86 * . . −0.600.18 Phe 230 A . . B . . . −1.43 0.86 * . . −0.60 0.17 Arg 231 A . . B .. . −0.62 0.61 * . . −0.60 0.21 Cys 232 . . . B T . . −0.37 0.53 * . .−0.20 0.41 Ile 233 . . . B T . . −0.27 0.46 * . . −0.20 0.46 Gln 234 . .. B T . . 0.54 0.10 * . . 0.10 0.37 Asn 235 . . . B . . C 0.93 0.10 * .. 0.05 1.19 Met 236 . . . B . . C 0.01 0.01 * . F 0.20 2.44 Pro 237 . .. B . . C 0.47 0.01 * . F 0.44 1.16 Glu 238 . . . . T . . 1.36 0.04 * .F 1.08 1.12 Thr 239 . . . . . . C 1.36 0.04 * . F 1.12 1.82 Leu 240 . .. . . . C 1.06 −0.17 * . F 1.96 1.89 Pro 241 . . . . T . . 0.99 −0.21 .. F 2.40 1.46 Asn 242 . . . . T . . 0.96 0.36 . . F 1.41 0.54 Asn 243 .. . . T T . 0.66 0.63 . . F 1.22 1.03 Ser 244 . . . . T T . 0.38 0.33 .. F 1.13 0.89 Cys 245 . . . . T T . 0.84 0.40 . . . 0.74 0.56 Tyr 246 .. . . T T . 0.17 0.43 . . . 0.20 0.35 Ser 247 A . . . . . . −0.42 0.71 .. . −0.40 0.18 Ala 248 A A . . . . . −0.38 0.83 . . . −0.60 0.34 Gly 249A A . . . . . −0.89 0.26 . . . −0.30 0.43 Ile 250 A A . . . . . −0.220.19 * . . −0.30 0.27 Ala 251 A A . . . . . 0.02 −0.20 * . . 0.30 0.46Lys 252 A A . . . . . −0.02 −0.70 . . . 0.60 0.80 Leu 253 A A . . . . .0.57 −0.70 . . F 0.90 1.13 Glu 254 A A . . . . . 0.91 −1.39 . . F 0.901.87 Glu 255 A A . . . . . 0.99 −1.89 . . F 0.90 1.62 Gly 256 A A . . .. . 1.58 −1.20 . * F 0.90 1.62 Asp 257 A A . . . . . 0.72 −1.49 . * F0.90 1.62 Glu 258 A A . . . . . 0.94 −0.80 * * F 0.75 0.77 Leu 259 A A .. . . . 0.06 −0.30 * * . 0.30 0.79 Gln 260 A A . . . . . −0.16 −0.04 * .. 0.30 0.33 Leu 261 A A . . . . . 0.30 0.39 * . . −0.30 0.30 Ala 262 A A. . . . . 0.30 0.39 * . . −0.30 0.70 Ile 263 A A . . . . . 0.30 −0.30. * . 0.30 0.70 Pro 264 A . . . . T . 0.52 −0.30 . * F 1.00 1.37 Arg 265A . . . . T . 0.52 −0.49 . * F 1.00 1.37 Glu 266 A . . . . T . 0.44−0.59 * * F 1.30 3.38 Asn 267 A . . . . T . 0.73 −0.59 * * F 1.30 1.53Ala 268 A . . . . . . 0.81 −0.63 * * . 0.95 1.05 Gln 269 A . . . . . .1.02 0.06 * * . −0.10 0.50 Ile 270 A . . . . . . 0.57 0.06 . * . 0.150.52 Ser 271 . . . . . . C 0.57 0.09 . * . 0.60 0.51 Leu 272 . . . . . .C −0.29 −0.41 . * F 1.60 0.49 Asp 273 . . . . T T . −0.01 −0.17 . * F2.25 0.52 Gly 274 . . . . T T . −0.71 −0.37 . * F 2.50 0.56 Asp 275 . .. . T T . −0.52 0.03 . * F 1.65 0.59 Val 276 A . . . . T . −0.57 0.13. * F 1.00 0.30 Thr 277 A . . B . . . −0.34 0.56 . * . −0.10 0.30 Phe278 A . . B . . . −1.16 0.63 . * . −0.35 0.18 Phe 279 A . . B . . .−0.77 1.31 . * . −0.60 0.20 Gly 280 A A . . . . . −1.58 0.67 . * . −0.600.28 Ala 281 A A . . . . . −1.53 0.87 . * . −0.60 0.27 Leu 282 A A . . .. . −1.61 0.77 * . . −0.60 0.26 Lys 283 A A . . . . . −1.30 0.41 * . .−0.60 0.33 Leu 284 A A . . . . . −0.99 0.41 . . . −0.60 0.42 Leu 285 A A. . . . . −1.03 0.34 * . . −0.30 0.65

TABLE 10 Res Position I II III IV V VI VII VIII IX X XI XII XIII XIV Met1 A . . . . . . 0.73 −0.71 . . . 0.95 1.39 Asp 2 A . . . . T . 1.12−0.66 * . . 1.15 1.56 Asp 3 A . . . . T . 1.62 −1.09 * . . 1.15 2.12 Ser4 A . . . . T . 2.01 −1.51 . . . 1.15 4.19 Thr 5 A . . . . T . 2.40−2.13 . . F 1.30 4.35 Glu 6 A A . . . . . 2.70 −1.73 * * F 0.90 4.51 Arg7 A A . . . . . 2.81 −1.34 * * F 0.90 4.51 Glu 8 A A . . . . . 2.00−1.73 * * F 0.90 6.12 Gln 9 A A . . . . . 1.99 −1.53 * * F 0.90 2.91 Ser10 A . . B . . . 2.00 −1.04 * * F 0.90 2.15 Arg 11 A . . B . . . 1.33−0.66 * * F 0.90 1.66 Leu 12 A . . B . . . 0.41 −0.09 * * F 0.45 0.51Thr 13 A . . B . . . 0.46 0.20 * * F −0.15 0.32 Ser 14 A A . . . . .0.50 −0.19 * * . 0.30 0.32 Cys 15 A A . . . . . 0.91 −0.19 * * . 0.300.78 Leu 16 A A . . . . . 0.80 −0.87 * * F 0.90 1.06 Lys 17 A A . . . .. 1.61 −1.36 . * F 0.90 1.37 Lys 18 A A . . . . . 1.32 −1.74 . * F 0.904.44 Arg 19 A A . . . . . 1.67 −1.70 . * F 0.90 5.33 Glu 20 A A . . . .. 1.52 −2.39 . * F 0.90 5.33 Glu 21 A A . . . . . 2.38 −1.70 . * F 0.902.20 Met 22 A A . . . . . 2.33 −1.70 . * F 0.90 2.24 Lys 23 A A . . . .. 1.62 −1.70 * * F 0.90 2.24 Leu 24 A A . . . . . 0.66 −1.13 * * F 0.750.69 Lys 25 A A . . . . . 0.36 −0.49 . * F 0.45 0.52 Glu 26 A A . B . .. −0.53 −0.71 * * . 0.60 0.35 Cys 27 A A . B . . . −0.74 −0.03 * * .0.30 0.30 Val 28 A A . B . . . −1.00 −0.03 * * . 0.30 0.12 Ser 29 A A .B . . . −0.08 0.40 * * . −0.30 0.11 Ile 30 A . . B . . . −0.08 0.40 * *. −0.30 0.40 Leu 31 A . . B . . . −0.08 −0.17 * . . 0.45 1.08 Pro 32 . .. B . . C 0.29 −0.81 * . F 1.10 1.39 Arg 33 . . . . T . . 0.93 −0.81 . *F 1.50 2.66 Lys 34 . . . . T . . 0.93 −1.07 . . F 1.84 4.98 Glu 35 . . .. . . C 0.97 −1.37 * * F 1.98 4.32 Ser 36 . . . . . T C 1.89 −1.16 * * F2.52 1.64 Pro 37 . . . . . T C 1.80 −1.16 * * F 2.86 1.60 Ser 38 . . . .T T . 1.39 −0.77 * . F 3.40 1.24 Val 39 A . . . . T . 1.39 −0.39 . * F2.36 1.24 Arg 40 A . . . . . . 1.39 −0.77 * * F 2.46 1.60 Ser 41 A . . .. . . 1.34 −1.20 * * F 2.46 2.00 Ser 42 . . . . T T . 1.60 −1.16 . * F3.06 2.67 Lys 43 . . . . T T . 1.09 −1.80 * * F 3.06 2.72 Asp 44 . . . .T T . 1.13 −1.11 * * F 3.40 1.67 Gly 45 A . . . . T . 0.43 −0.81 * * F2.66 1.03 Lys 46 A A . . . . . 0.14 −0.70 . . F 1.77 0.52 Leu 47 A A . .. . . 0.13 −0.20 * . . 0.98 0.31 Leu 48 A A . . . . . −0.72 0.29 * . .0.04 0.46 Ala 49 A A . . . . . −1.53 0.54 . * . −0.60 0.19 Ala 50 A A .. . . . −2.00 1.23 . . . −0.60 0.19 Thr 51 A A . . . . . −2.63 1.23 . .. −0.60 0.19 Leu 52 A A . . . . . −2.63 1.04 . . . −0.60 0.19 Leu 53 A A. . . . . −2.63 1.23 . . . −0.60 0.15 Leu 54 A A . . . . . −2.34 1.41 .. . −0.60 0.09 Ala 55 A A . . . . . −2.42 1.31 . . . −0.60 0.14 Leu 56 AA . . . . . −2.78 1.20 . . . −0.60 0.09 Leu 57 A . . . . T . −2.78 1.09. . . −0.20 0.06 Ser 58 A . . . . T . −2.28 1.09 . . . −0.20 0.05 Cys 59A . . . . T . −2.32 1.07 . . . −0.20 0.09 Cys 60 A . . . . T . −2.591.03 . . . −0.20 0.08 Leu 61 . . B B . . . −2.08 0.99 . . . −0.60 0.04Thr 62 . . B B . . . −1.97 0.99 . . . −0.60 0.11 Val 63 . . B B . . .−1.91 1.20 . . . −0.60 0.17 Val 64 . . B B . . . −1.24 1.39 . . . −0.600.33 Ser 65 . . B B . . . −1.43 1.10 . . . −0.60 0.40 Phe 66 A . . B . .. −1.21 1.26 . . . −0.60 0.40 Tyr 67 A . . B . . . −1.49 1.11 . . .−0.60 0.54 Gln 68 A . . B . . . −1.44 0.97 . . . −0.60 0.41 Val 69 A . .B . . . −0.59 1.27 . . . −0.60 0.39 Ala 70 A . . B . . . −0.63 0.89 . .. −0.60 0.43 Ala 71 A . . B . . . 0.07 0.56 . * . −0.60 0.25 Leu 72 A .. . . T . −0.50 0.16 . . . 0.10 0.55 Gln 73 A . . . . T . −1.09 0.20 . .F 0.25 0.45 Gly 74 A . . . . T . −0.53 0.20 . . F 0.25 0.45 Asp 75 A . .. . T . −0.76 0.09 . * F 0.25 0.73 Leu 76 A A . . . . . −0.06 0.09 . * F−0.15 0.35 Ala 77 A A . . . . . 0.17 −0.31 . * . 0.30 0.69 Ser 78 A A .. . . . 0.17 −0.24 . * . 0.30 0.42 Leu 79 A A . . . . . −0.30 −0.24 . *. 0.30 0.88 Arg 80 A A . . . . . −0.30 −0.24 . * . 0.30 0.72 Ala 81 A A. . . . . 0.17 −0.34 . * . 0.30 0.93 Glu 82 A A . . . . . 0.72 −0.30 . *. 0.45 1.11 Leu 83 A A . . . . . 0.99 −0.49 . * . 0.30 0.77 Gln 84 A A .. . . . 1.21 0.01 . * . −0.15 1.04 Gly 85 A A . . . . . 1.10 0.01 * * .−0.30 0.61 His 86 A A . . . . . 1.73 0.01 * * . −0.15 1.27 His 87 A A .. . . . 0.92 −0.67 . * . 0.75 1.47 Ala 88 A A . . . . . 1.52 −0.39 . * .0.45 1.22 Glu 89 A A . . . . . 0.93 −0.39 . . . 0.45 1.39 Lys 90 A A . .. . . 0.93 −0.39 * . F 0.60 1.03 Leu 91 A . . . . T . 0.38 −0.46 * . .0.85 1.01 Pro 92 A . . . . T . 0.07 −0.46 . . . 0.70 0.59 Ala 93 A . . .. T . 0.07 −0.03 . . . 0.70 0.29 Gly 94 A . . . . T . −0.14 0.47 . . .−0.20 0.36 Ala 95 A . . . . . . −0.14 0.21 . * . −0.10 0.36 Gly 96 A . .. . . . 0.08 −0.21 . . F 0.65 0.71 Ala 97 A . . . . . . −0.06 −0.21 . .F 0.65 0.72 Pro 98 A . . . . . . −0.28 −0.21 . * F 0.65 0.71 Lys 99 A A. . . . . 0.07 −0.03 . . F 0.45 0.59 Ala 100 A A . . . . . 0.66 −0.46 .. F 0.60 1.01 Gly 101 A A . . . . . 0.41 −0.96 . . F 0.90 1.13 Leu 102 AA . . . . . 0.79 −0.89 . . F 0.75 0.57 Glu 103 A A . . . . . 0.41−0.46 * . F 0.45 0.88 Glu 104 A A . . . . . −0.49 −0.46 * . F 0.45 0.89Ala 105 A A . . . . . −0.21 −0.24 . . . 0.30 0.81 Pro 106 A A . . . . .−0.46 −0.44 . . . 0.30 0.67 Ala 107 A A . . . . . 0.01 0.06 . . . −0.300.39 Val 108 A A . . . . . −0.80 0.49 * * . −0.60 0.38 Thr 109 A A . . .. . −0.76 0.67 . * . −0.60 0.20 Ala 110 A A . . . . . −1.06 0.24 * * .−0.30 0.40 Gly 111 A A . . . . . −1.54 0.43 * * . −0.60 0.38 Leu 112 A A. . . . . −0.96 0.57 * * . −0.60 0.23 Lys 113 . A B . . . . −0.310.09 * * . −0.30 0.39 Ile 114 . A B . . . . −0.21 0.01 * . . −0.30 0.61Phe 115 . A B . . . . −0.21 0.01 * . . 0.15 1.15 Glu 116 . A . . . . C−0.08 −0.17 * . F 1.25 0.58 Pro 117 . A . . . . C 0.39 0.26 * * F 1.101.28 Pro 118 . . . . . . C 0.34 0.00 * . F 2.20 1.47 Ala 119 . . . . . TC 0.89 −0.79 . * F 3.00 1.47 Pro 120 . . . . . T C 1.59 −0.36 . * F 2.250.94 Gly 121 . . . . T T . 1.29 −0.39 . * F 2.15 0.98 Glu 122 . . . . TT . 1.20 −0.43 . . F 2.00 1.30 Gly 123 . . . . . . C 1.41 −0.54 . . F1.60 1.12 Asn 124 . . . . . T C 2.00 −0.57 . . F 1.50 1.97 Ser 125 . . .. . T C 1.91 −0.60 . * F 1.50 1.82 Ser 126 . . . . . T C 2.37 −0.21 . *F 1.54 2.47 Gln 127 . . . . . T C 2.37 −0.64 . * F 2.18 3.01 Asn 128 . .. . . . C 2.76 −0.64 . . F 2.32 3.61 Ser 129 . . . . . T C 2.87 −1.03 .. F 2.86 5.39 Arg 130 . . . . T T . 2.58 −1.41 * . F 3.40 6.09 Asn 131 .. . . T T . 2.02 −1.31 * . F 3.06 3.83 Lys 132 . . . . T T . 2.02−1.07 * . F 2.72 2.12 Arg 133 . . . . T . . 1.68 −1.06 * . F 2.18 1.88Ala 134 . . . . . . C 1.77 −0.63 * . F 1.64 1.15 Val 135 . . . . . . C1.66 −0.60 * . F 1.15 0.89 Gln 136 . . . . . . C 1.66 −0.60 * . F 1.490.79 Gly 137 . . . . . T C 1.30 −0.60 * . F 2.18 1.35 Pro 138 . . . . .T C 0.84 −0.61 * . F 2.52 2.63 Glu 139 . . . . . T C 1.13 −0.83 * . F2.86 1.50 Glu 140 . . . . T T . 1.74 −0.84 . . F 3.40 2.03 Thr 141 . . .. T . . 1.43 −0.51 . . F 2.86 2.06 Gly 142 . . . . T T . 1.08 −0.46 . .F 2.42 1.72 Ser 143 . . . . T T . 0.43 0.33 . . F 1.33 0.86 Tyr 144 . .. . T T . 0.22 0.97 . . . 0.54 0.44 Thr 145 . . . . T T . −0.07 0.91 . .. 0.20 0.69 Phe 146 . . B B . . . −0.57 1.40 . . . −0.60 0.54 Val 147 .. B B . . . −1.03 1.70 . . . −0.60 0.29 Pro 148 . . B B . . . −1.03 1.63. . . −0.60 0.16 Trp 149 A . . B . . . −1.49 1.53 . * . −0.60 0.25 Leu150 A . . B . . . −1.13 1.53 * . . −0.60 0.29 Leu 151 A . . B . . .−0.32 0.89 * . . −0.30 0.38 Ser 152 A . . . . . . 0.19 0.46 * . . 0.200.71 Phe 153 . . . . T . . 0.10 −0.03 * . . 1.80 0.85 Lys 154 . . . . TT . −0.20 −0.33 * . F 2.60 1.38 Arg 155 . . . . . T C −0.20 −0.51 . . F3.00 1.04 Gly 156 . . . . . T C 0.61 −0.21 . . F 2.25 0.99 Ser 157 A . .. . T . 0.91 −1.00 * . F 2.05 0.86 Ala 158 A A . . . . . 1.66 −1.00 * .F 1.35 0.76 Leu 159 A A . . . . . 1.61 −1.00 . . F 1.20 1.54 Glu 160 A A. . . . . 1.50 −1.43 . . F 0.90 1.98 Glu 161 A A . . . . . 1.89 −1.41 *. F 0.90 3.16 Lys 162 A A . . . . . 1.30 −1.91 * . F 0.90 7.66 Glu 163 AA . . . . . 1.08 −1.91 . . F 0.90 3.10 Asn 164 A A . . . . . 1.03−1.23 * * F 0.90 1.48 Lys 165 A A . . . . . 1.08 −0.59 * . F 0.75 0.55Ile 166 A A . . . . . 1.08 −0.59 * * . 0.60 0.63 Leu 167 A A . . . . .0.72 −0.59 * * . 0.76 0.68 Val 168 A A . . . . . 0.38 −0.50 . * . 0.920.49 Lys 169 A A . . . . . 0.13 −0.07 * * F 0.93 0.69 Glu 170 A . . . .T . −0.61 0.00 * * F 1.64 1.32 Thr 171 . . . . T T . −0.42 0.10 . * F1.60 1.54 Gly 172 . . . . T T . −0.50 0.24 * . F 1.29 0.67 Tyr 173 . . .. T T . 0.11 0.93 * * . 0.68 0.27 Phe 174 . . B B . . . −0.28 1.69 . . .−0.28 0.29 Phe 175 . . B B . . . −0.28 1.63 . * . −0.44 0.29 Ile 176 . .B B . . . −0.82 1.60 . . . −0.60 0.32 Tyr 177 . . B B . . . −1.29 1.49 .. . −0.60 0.28 Gly 178 . . . B T . . −1.29 1.39 . . . −0.20 0.26 Gln 179. . . B T . . −0.90 1.36 . . . −0.20 0.59 Val 180 . . . B . . C −0.201.16 . . . −0.40 0.54 Leu 181 . . . B . . C 0.73 0.40 . . . −0.10 0.92Tyr 182 . . . . T T . 0.67 −0.03 . . . 1.25 1.06 Thr 183 . . . . T T .0.77 0.06 . . F 0.80 2.06 Asp 184 . . . . T T . 0.18 0.17 . . F 0.803.91 Lys 185 A . . . . T . 0.43 −0.01 . . F 1.00 2.52 Thr 186 A A . . .. . 0.90 −0.16 . . F 0.60 1.73 Tyr 187 A A . . . . . 1.11 −0.21 . . .0.45 1.03 Ala 188 A A . . . . . 0.61 0.29 . . . −0.30 0.70 Met 189 A A .. . . . −0.28 0.97 . . . −0.60 0.40 Gly 190 A A . B . . . −0.32 1.17 * .. −0.60 0.18 His 191 A A . B . . . 0.10 0.81 * . . −0.60 0.31 Leu 192 AA . B . . . 0.39 0.31 . . . −0.30 0.61 Ile 193 A A . B . . . 1.02 −0.30. . . 0.45 1.22 Gln 194 A A . B . . . 0.77 −0.73 . * . 0.75 1.80 Arg 195A A . B . . . 1.08 −0.59 * * F 0.90 1.62 Lys 196 A A . B . . . 0.26−0.77 * * F 0.90 3.14 Lys 197 A A . B . . . 0.37 −0.81 . * F 0.90 1.35Val 198 . A B B . . . 0.91 −0.43 * * . 0.30 0.60 His 199 . A B B . . .0.91 0.00 * * . 0.30 0.29 Val 200 . A B B . . . 0.80 0.00 * * . 0.300.25 Phe 201 . . B B . . . −0.06 0.00 * . . 0.30 0.57 Gly 202 A . . B .. . −0.40 0.04 . * . −0.30 0.35 Asp 203 A . . . . . . −0.36 −0.07 * . .0.50 0.63 Glu 204 A . . . . . . −1.18 −0.03 * . . 0.50 0.60 Leu 205 A .. B . . . −0.63 −0.17 . . . 0.30 0.45 Ser 206 A . . B . . . −0.74 −0.11. . . 0.30 0.39 Leu 207 A . . B . . . −1.10 0.57 . * . −0.60 0.18 Val208 A . . B . . . −0.99 1.36 . * . −0.60 0.19 Thr 209 A . . B . . .−1.66 0.67 * * . −0.60 0.28 Leu 210 A . . B . . . −1.73 0.86 * . . −0.600.18 Phe 211 A . . B . . . −1.43 0.86 * . . −0.60 0.17 Arg 212 A . . B .. . −0.62 0.61 * . . −0.60 0.21 Cys 213 . . . B T . . −0.37 0.53 * . .−0.20 0.41 Ile 214 . . . B T . . −0.27 0.46 * . . −0.20 0.46 Gln 215 . .. B T . . 0.54 0.10 * . . 0.10 0.37 Asn 216 . . . B . . C 0.93 0.10 * .. 0.05 1.19 Met 217 . . . B . . C 0.01 0.01 * . F 0.20 2.44 Pro 218 . .. B . . C 0.47 0.01 * . F 0.44 1.16 Glu 219 . . . . T . . 1.36 0.04 * .F 1.08 1.12 Thr 220 . . . . . . C 1.36 0.04 * . F 1.12 1.82 Leu 221 . .. . . . C 1.06 −0.17 * . F 1.96 1.89 Pro 222 . . . . T . . 0.99 −0.21 .. F 2.40 1.46 Asn 223 . . . . T . . 0.96 0.36 . . F 1.41 0.54 Asn 224 .. . . T T . 0.66 0.63 . . F 1.22 1.03 Ser 225 . . . . T T . 0.38 0.33 .. F 1.13 0.89 Cys 226 . . . . T T . 0.84 0.40 . . . 0.74 0.56 Tyr 227 .. . . T T . 0.17 0.43 . . . 0.20 0.35 Ser 228 A . . . . . . −0.42 0.71 .. . −0.40 0.18 Ala 229 A A . . . . . −0.38 0.83 . . . −0.60 0.34 Gly 230A A . . . . . −0.89 0.26 . . . −0.30 0.43 Ile 231 A A . . . . . −0.220.19 * . . −0.30 0.27 Ala 232 A A . . . . . 0.02 −0.20 * . . 0.30 0.46Lys 233 A A . . . . . −0.02 −0.70 . . . 0.60 0.80 Leu 234 A A . . . . .0.57 −0.70 . . F 0.90 1.13 Glu 235 A A . . . . . 0.91 −1.39 . . F 0.901.87 Glu 236 A A . . . . . 0.99 −1.89 . . F 0.90 1.62 Gly 237 A A . . .. . 1.58 −1.20 . * F 0.90 1.62 Asp 238 A A . . . . . 0.72 −1.49 . * F0.90 1.62 Glu 239 A A . . . . . 0.94 −0.80 * * F 0.75 0.77 Leu 240 A A .. . . . 0.06 −0.30 * * . 0.30 0.79 Gln 241 A A . . . . . −0.16 −0.04 * .. 0.30 0.33 Leu 242 A A . . . . . 0.30 0.39 * . . −0.30 0.30 Ala 243 A A. . . . . 0.30 0.39 * . . −0.30 0.70 Ile 244 A A . . . . . 0.30 −0.30. * . 0.30 0.70 Pro 245 A . . . . T . 0.52 −0.30 . * F 1.00 1.37 Arg 246A . . . . T . 0.52 −0.49 . * F 1.00 1.37 Glu 247 A . . . . T . 0.44−0.59 * * F 1.30 3.38 Asn 248 A . . . . T . 0.73 −0.59 * * F 1.30 1.53Ala 249 A . . . . . . 0.81 −0.63 * * . 0.95 1.05 Gln 250 A . . . . . .1.02 0.06 * * . −0.10 0.50 Ile 251 A . . . . . . 0.57 0.06 * * . 0.150.52 Ser 252 . . . . . . C 0.57 0.09 . * . 0.60 0.51 Leu 253 . . . . . .C −0.29 −0.41 . * F 1.60 0.49 Asp 254 . . . . T T . −0.01 −0.17 . * F2.25 0.52 Gly 255 . . . . T T . −0.71 −0.37 . * F 2.50 0.56 Asp 256 . .. . T T . −0.52 0.03 . * F 1.65 0.59 Val 257 A . . . . T . −0.57 0.13. * F 1.00 0.30 Thr 258 A . . B . . . −0.34 0.56 . * . −0.10 0.30 Phe259 A . . B . . . −1.16 0.63 . * . −0.35 0.18 Phe 260 A . . B . . .−0.77 1.31 . * . −0.60 0.20 Gly 261 A A . . . . . −1.58 0.67 . * . −0.600.28 Ala 262 A A . . . . . −1.53 0.87 . * . −0.60 0.27 Leu 263 A A . . .. . −1.61 0.77 * . . −0.60 0.26 Lys 264 A A . . . . . −1.30 0.41 * . .−0.60 0.33 Leu 265 A A . . . . . −0.99 0.41 . . . −0.60 0.42 Leu 266 A A. . . . . −1.03 0.34 * . . −0.30 0.65

In another embodiment, the invention provides antibodies that bind apolypeptide comprising, or alternatively consisting of, anepitope-bearing portion of a polypeptide of the invention. The epitopeof this polypeptide portion may be an immunogenic or antigenic epitopeof a polypeptide of the invention. An “immunogenic epitope” is definedas a part of a protein that elicits an antibody response when the wholeprotein is the immunogen. On the other hand, a region of a proteinmolecule to which an antibody can bind is defined as an “antigenicepitope.” The number of immunogenic epitopes of a protein generally isless than the number of antigenic epitopes. See, for instance, Geysen etal., Proc. Natl. Acad. Sci. USA 81:3998-4002 (1983).

As to the selection of polypeptides bearing an antigenic epitope (i.e.,that contain a region of a protein molecule to which an antibody canbind), it is well known in that art that relatively short syntheticpeptides that mimic part of a protein sequence are routinely capable ofeliciting an antiserum that reacts with the partially mimicked protein.See, for instance, Sutcliffe, J. G., Shinnick, T. M., Green, N. andLearner, R. A. (1983) “Antibodies that react with predetermined sites onproteins”, Science, 219:660-666. Peptides capable of elicitingprotein-reactive sera are frequently represented in the primary sequenceof a protein, can be characterized by a set of simple chemical rules,and are confined neither to immunodominant regions of intact proteins(i.e., immunogenic epitopes) nor to the amino or carboxyl terminals.Antigenic epitope-bearing peptides and polypeptides of the invention aretherefore useful to raise antibodies, including monoclonal antibodies,that bind specifically to a polypeptide of the invention. See, forinstance, Wilson et al., Cell 37:767-778 (1984) at 777.

In specific embodiments, antibodies of the present invention bindantigenic epitope-bearing peptides and polypeptides of B LymphocyteStimulator and preferably contain a sequence of at least 4, at least 5,at least 6, at least 7, more preferably at least 8, at least 9, at least10, at least 11, at least 12, at least 13, at least 14, at least 15, atleast 20, at least 25, at least 30, at least 40, at least 50, and, mostpreferably, between about 15 to about 30 amino acids contained withinthe amino acid sequence of a B Lymphocyte Stimulator polypeptide.Preferred polypeptides comprising immunogenic or antigenic epitopes areat least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85,90, 95, or 100 amino acid residues in length. Additional non-exclusivepreferred antigenic epitopes include the antigenic epitopes disclosedherein, as well as portions thereof.

Non-limiting examples of antigenic polypeptides or peptides that can beused to generate B Lymphocyte Stimulator-specific antibodies and whichmay be bound by the antibodies of the invention include: a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Phe-115 to about Leu-147 in SEQ ID NO:3228; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Ile-150 to about Tyr-163 in SEQ ID NO:3228; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Ser-171 to about Phe-194 in SEQ ID NO:3228; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Glu-223 to about Tyr-246 in SEQ ID NO:3228; and a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Ser-271 to about Phe-278 in FIGS. 1A and 1B (SEQ ID NO:3228). Inthis context, “about” means the particularly recited ranges and rangeslarger or smaller by several, a few, 5, 4, 3, 2 or 1 amino acid residuesat either or both the amino- and carboxy-termini. These polypeptidefragments have been determined to bear antigenic epitopes of the BLymphocyte Stimulator polypeptide by the analysis of the Jameson-Wolfantigenic index, as disclosed Table 9, above.

Non-limiting examples of antigenic polypeptides or peptides that can beused to generate B Lymphocyte Stimulator-specific antibodies and whichmay be bound by the antibodies of the invention include: a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Pro-32 to about Leu-47 in SEQ ID NO:3229; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Glu-116 to about Ser-143 in SEQ ID NO:3229; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Phe-153 to about Tyr-173 in SEQ ID NO:3229; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Pro-218 to about Tyr-227 in SEQ ID NO:3229; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Ala-232 to about Gln-241 in SEQ ID NO:3229; a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Ile-244 to about Ala-249 in SEQ ID NO:3229; and a polypeptidecomprising, or alternatively consisting of, amino acid residues fromabout Ser-252 to about Val-257 in SEQ ID NO:3229. In this context,“about” means the particularly recited ranges and ranges larger orsmaller by several, a few, 5, 4, 3, 2 or 1 amino acid residues at eitheror both the amino- and carboxy-termini. These polypeptide fragments havebeen determined to bear antigenic epitopes of the B LymphocyteStimulator polypeptide by the analysis of the Jameson-Wolf antigenicindex, as disclosed in Table 10 generated by the Protean component ofthe DNA*STAR computer program (as set forth above).

B Lymphocyte Stimulator epitope-bearing peptides and polypeptides may beproduced by any conventional means. See, e.g., Houghten, R. A. (1985)General method for the rapid solid-phase synthesis of large numbers ofpeptides: specificity of antigen-antibody interaction at the level ofindividual amino acids. Proc. Natl. Acad. Sci. USA 82:5131-5135; this“Simultaneous Multiple Peptide Synthesis (SMPS)” process is furtherdescribed in U.S. Pat. No. 4,631,211 to Houghten et al. (1986).

The present invention encompasses antibodies that bind polypeptidescomprising, or alternatively consisting of, an epitope of thepolypeptide having an amino acid sequence of SEQ ID NO:3228, or anepitope of the polypeptide sequence encoded by a polynucleotide sequencecontained in ATCC™ deposit No. 97768, or encoded by a polynucleotidethat hybridizes to cDNA sequence contained in ATCC™ deposit No. 97768(e.g., under hybridization conditions described herein).

The present invention also encompasses antibodies that bind polypeptidescomprising, or alternatively consisting of, an epitope of thepolypeptide having an amino acid sequence of SEQ ID NO:3229, or anepitope of the polypeptide sequence encoded by a polynucleotide sequencecontained in ATCC™ deposit No. 203518, or encoded by a polynucleotidethat hybridizes to the cDNA sequence contained in ATCC™ deposit No.203518 (e.g., under hybridization conditions described herein).

The term “epitopes,” as used herein, refers to portions of a polypeptidehaving antigenic or immunogenic activity in an animal, preferably amammal, and most preferably in a human. In a preferred embodiment, thepresent invention encompasses antibodies that bind a polypeptidecomprising an epitope. An “immunogenic epitope,” as used herein, isdefined as a portion of a protein that elicits an antibody response inan animal, as determined by any method known in the art, for example, bythe methods for generating antibodies described infra. (See, forexample, Geysen et al., Proc. Natl. Acad. Sci. USA 81:3998-4002 (1983)).The term “antigenic epitope,” as used herein, is defined as a portion ofa protein to which an antibody can immunospecifically bind its antigenas determined by any method well known in the art, for example, by theimmunoassays described herein Immunospecific binding excludesnon-specific binding but does not necessarily exclude cross-reactivitywith other antigens. Antigenic epitopes need not necessarily beimmunogenic.

B Lymphocyte Stimulator polypeptide fragments which function as epitopesmay be produced by any conventional means. (See, e.g., Houghten, Proc.Natl. Acad. Sci. USA 82:5131-5135 (1985), further described in U.S. Pat.No. 4,631,211).

In the present invention, antibodies of the present invention bindantigenic epitopes preferably containing a sequence of at least 4, atleast 5, at least 6, at least 7, more preferably at least 8, at least 9,at least 10, at least 11, at least 12, at least 13, at least 14, atleast 15, at least 20, at least 25, at least 30, at least 40, at least50, and, most preferably, between about 15 to about 30 amino acids.Preferred polypeptides comprising immunogenic or antigenic epitopes thatmay be bound by antibodies of the present invention are at least 10, 15,20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100amino acid residues in length. Additional non-exclusive preferredantigenic epitopes include the antigenic epitopes disclosed herein, aswell as portions thereof. Antigenic epitopes are useful, for example, toraise antibodies, including monoclonal antibodies, that specificallybind the epitope. Preferred antigenic epitopes include the antigenicepitopes disclosed herein, as well as any combination of two, three,four, five or more of these antigenic epitopes. Antigenic epitopes canbe used as the target molecules in immunoassays. (See, for instance,Wilson et al., Cell 37:767-778 (1984); Sutcliffe et al., Science219:660-666 (1983)).

Similarly, immunogenic epitopes can be used, for example, to induceantibodies according to methods well known in the art. (See, forinstance, Sutcliffe et al., supra; Wilson et al., supra; Chow et al.,Proc. Natl. Acad. Sci. USA 82:910-914; and Bittle et al., J. Gen. Virol.66:2347-2354 (1985). Preferred immunogenic epitopes include theimmunogenic epitopes disclosed herein, as well as any combination oftwo, three, four, five or more of these immunogenic epitopes. Thepolypeptides comprising one or more immunogenic epitopes of B LymphocyteStimulator may be presented for eliciting an antibody response togetherwith a carrier protein, such as an albumin, to an animal system (such asrabbit or mouse), or, if the polypeptide is of sufficient length (atleast about 25 amino acids), the polypeptide may be presented without acarrier. However, immunogenic epitopes comprising as few as 8 to 10amino acids have been shown to be sufficient to raise antibodies capableof binding to, at the very least, linear epitopes in a denaturedpolypeptide (e.g., in Western blotting).

Epitope-bearing B Lymphocyte Stimulator polypeptides may be used toinduce antibodies according to methods well known in the art including,but not limited to, in vivo immunization, in vitro immunization, andphage display methods. See, e.g., Sutcliffe et al., supra; Wilson etal., supra, and Bittle et al., J. Gen. Virol., 66:2347-2354 (1985). Ifin vivo immunization is used, animals may be immunized with freepeptide; however, anti-peptide antibody titer may be boosted by couplingthe peptide to a macromolecular carrier, such as keyhole limpethemocyanin (KLH) or tetanus toxoid. For instance, peptides containingcysteine residues may be coupled to a carrier using a linker such asmaleimidobenzoyl-N-hydroxysuccinimide ester (MBS), while other peptidesmay be coupled to carriers using a more general linking agent such asglutaraldehyde Animals such as rabbits, rats and mice are immunized witheither free or carrier-coupled peptides, for instance, byintraperitoneal and/or intradermal injection of emulsions containingabout 100 micrograms of peptide or carrier protein and Freund's adjuvantor any other adjuvant known for stimulating an immune response. Severalbooster injections may be needed, for instance, at intervals of abouttwo weeks, to provide a useful titer of anti-peptide antibody which canbe detected, for example, by ELISA assay using free peptide adsorbed toa solid surface. The titer of anti-peptide antibodies in serum from animmunized animal may be increased by selection of anti-peptideantibodies, for instance, by adsorption to the peptide on a solidsupport and elution of the selected antibodies according to methods wellknown in the art.

As one of skill in the art will appreciate, and as discussed above, theantibodies of the present invention may bind polypeptides comprising animmunogenic or antigenic epitope fused to other polypeptide sequences.For example, the B Lymphocyte Stimulator polypeptides may be fused withthe constant domain of immunoglobulins (IgA, IgE, IgG, IgM), or portionsthereof (CH1, CH2, CH3, or any combination thereof and portionsthereof), or albumin (including but not limited to recombinant humanalbumin or fragments or variants thereof (see, e.g., U.S. Pat. No.5,876,969, issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No.5,766,883, issued Jun. 16, 1998, herein incorporated by reference intheir entirety)), resulting in chimeric polypeptides. Such fusionproteins may facilitate purification and may increase half-life in vivo.This has been shown for chimeric proteins consisting of the first twodomains of the human CD4-polypeptide and various domains of the constantregions of the heavy or light chains of mammalian immunoglobulins. See,e.g., EP 394,827; Traunecker et al., Nature, 331:84-86 (1988) Enhanceddelivery of an antigen across the epithelial barrier to the immunesystem has been demonstrated for antigens (e.g., insulin) conjugated toan FcRn binding partner such as IgG or Fc fragments (see, e.g., PCTPublications WO 96/22024 and WO 99/04813). IgG Fusion proteins that havea disulfide-linked dimeric structure due to the IgG portion disulfidebonds have also been found to be more efficient in binding andneutralizing other molecules than monomeric polypeptides or fragmentsthereof alone. See, e.g., Fountoulakis et al., J. Biochem.,270:3958-3964 (1995). Nucleic acids encoding the above epitopes can alsobe recombined with a gene of interest as an epitope tag (e.g., thehemagglutinin (“HA”) tag or flag tag) to aid in detection andpurification of the expressed polypeptide. For example, a systemdescribed by Janknecht et al. allows for the ready purification ofnon-denatured fusion proteins expressed in human cell lines (Janknechtet al., 1991, Proc. Natl. Acad. Sci. USA 88:8972-897). In this system,the gene of interest is subcloned into a vaccinia recombination plasmidsuch that the open reading frame of the gene is translationally fused toan amino-terminal tag consisting of six histidine residues. The tagserves as a matrix-binding domain for the fusion protein. Extracts fromcells infected with the recombinant vaccinia virus are loaded onto Ni²⁺nitriloacetic acid-agarose column and histidine-tagged proteins can beselectively eluted with imidazole-containing buffers.

In another embodiment, the antibodies of the present invention bind BLymphocyte Stimulator polypeptides and/or the epitope-bearing fragmentsthereof that are fused with a heterologous antigen (e.g., polypeptide,carbohydrate, phospholipid, or nucleic acid). In specific embodiments,the heterologous antigen is an immunogen.

In a more specific embodiment, the heterologous antigen is the gp120protein of HIV, or a fragment thereof.

In another embodiment, antibodies of the present invention bind BLymphocyte Stimulator polypeptides and/or the epitope-bearing fragmentsthereof that are fused with polypeptide sequences of another TNF ligandfamily member (or biologically active fragments or variants thereof). Ina specific embodiment, the antibodies of the present invention bind BLymphocyte Stimulator polypeptides of the present invention are fusedwith a CD40L polypeptide sequence. In a preferred embodiment, the CD40Lpolypeptide sequence is soluble.

In another embodiment, antibodies of the present invention bind mutant BLymphocyte Stimulator polypeptides that have been generated by randommutagenesis of a polynucleotide encoding the B Lymphocyte Stimulatorpolypeptide, by error-prone PCR, random nucleotide insertion or othermethods prior to recombination. In another embodiment, antibodies of thepresent invention bind one or more components, motifs, sections, parts,domains, fragments, etc., of B Lymphocyte Stimulator recombined with oneor more components, motifs, sections, parts, domains, fragments, etc. ofone or more heterologous molecules. In preferred embodiments, theheterologous molecules are, for example, TNF-alpha, lymphotoxin-alpha(LT-alpha, also known as TNF-beta), LT-beta (found in complexheterotrimer LT-alpha2-beta), OPGL, FasL, CD27L, CD30L, CD40L, 4-1BBL,DcR3, OX40L, TNF-gamma (International Publication No. WO 96/14328),AIM-I (International Publication No. WO 97/33899), AIM-II (InternationalPublication No. WO 97/34911), APRIL (J. Exp. Med. 188(6):1185-1190),endokine-alpha (International Publication No. WO 98/07880), OPG, OX40,and nerve growth factor (NGF), and soluble forms of Fas, CD30, CD27,CD40 and 4-IBB, TR2 (International Publication No. WO 96/34095), DR3(International Publication No. WO 97/33904), DR4 (InternationalPublication No. WO 98/32856), TR5 (International Publication No. WO98/30693), TR6 (International Publication No. WO 98/30694), TR7(International Publication No. WO 98/41629), TRANK, TR9 (InternationalPublication No. WO 98/56892), TR10 (International Publication No. WO98/54202), 312C2 (International Publication No. WO 98/06842), TR12, CAD,and v-FLIP. In further embodiments, the heterologous molecules are anymember of the TNF family.

In another preferred embodiment, antibodies of the present inventionbind B Lymphocyte Stimulator polypeptides of the invention (includingbiologically active fragments or variants thereof), that are fused withsoluble APRIL polypeptides (e.g., amino acid residues 105 through 250 ofSEQ ID NO:3239), or biologically active fragments or variants thereof.

To improve or alter the characteristics of B Lymphocyte Stimulatorpolypeptides, protein engineering may be employed. Recombinant DNAtechnology known to those skilled in the art can be used to create novelmutant proteins or “muteins including single or multiple amino acidsubstitutions, deletions, additions or fusion proteins. Such modifiedpolypeptides can show, e.g., enhanced activity or increased stability.In addition, they may be purified in higher yields and show bettersolubility than the corresponding natural polypeptide, at least undercertain purification and storage conditions. For instance, for manyproteins, including the extracellular domain or the mature form(s) of asecreted protein, it is known in the art that one or more amino acidsmay be deleted from the N-terminus or C-terminus without substantialloss of biological function. For instance, Ron et al., J. Biol. Chem.,268:2984-2988 (1993) reported modified KGF proteins that had heparinbinding activity even if 3, 8, or 27 amino-terminal amino acid residueswere missing. Accordingly, antibodies of the present invention may bindB Lymphocyte Stimulator polypeptide mutants or variants generated byprotein engineering.

In the present case, since the protein of the invention is a member ofthe TNF polypeptide family, deletions of N-terminal amino acids up tothe Gly (G) residue at position 191 in SEQ ID NO:3228 may retain somebiological activity such as, for example, the ability to stimulatelymphocyte (e.g., B cell) proliferation, differentiation, and/oractivation, and cytotoxicity to appropriate target cells. Polypeptideshaving further N-terminal deletions including the Gly (G) residue wouldnot be expected to retain biological activities because it is known thatthis residue in TNF-related polypeptides is in the beginning of theconserved domain required for biological activities. However, even ifdeletion of one or more amino acids from the N-terminus of a proteinresults in modification or loss of one or more biological functions ofthe protein, other functional activities may still be retained. Thus,the ability of the shortened protein to induce and/or bind to antibodieswhich recognize the complete or extracellular domain of the proteingenerally will be retained when less than the majority of the residuesof the complete or extracellular domain of the protein are removed fromthe N-terminus. Whether a particular polypeptide lacking N-terminalresidues of a complete protein retains such immunologic activities canreadily be determined by routine methods described herein and otherwiseknown in the art.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues deleted from the amino terminusof the amino acid sequence of the B Lymphocyte Stimulator of SEQ IDNO:3228, up to the glycine residue at position 191 (Gly-191 residue fromthe amino terminus). In particular, the present invention providesantibodies that bind polypeptides comprising, or alternativelyconsisting of, the amino acid sequence of residues n¹-285 of SEQ IDNO:3228, where n¹ is an integer in the range of the amino acid positionof amino acid residues 2-190 of the amino acid sequence in SEQ IDNO:3228. More in particular, the invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues 2-285, 3-285,4-285, 5-285, 6-285, 7-285, 8-285, 9-285, 10-285, 11-285, 12-285,13-285, 14-285, 15-285, 16-285, 17-285, 18-285, 19-285, 20-285, 21-285,22-285, 23-285, 24-285, 25-285, 26-285, 27-285, 28-285, 29-285, 30-285,31-285, 32-285, 33-285, 34-285, 35-285, 36-285, 37-285, 38-285, 39-285,40-285, 41-285, 42-285, 43-285, 44-285, 45-285, 46-285, 47-285, 48-285,49-285, 50-285, 51-285, 52-285, 53-285, 54-285, 55-285, 56-285, 57-285,58-285, 59-285, 60-285, 61-285, 62-285, 63-285, 64-285, 65-285, 66-285,67-285, 68-285, 69-285, 70-285, 71-285, 72-285, 73-285, 74-285, 75-285,76-285, 77-285, 78-285, 79-285, 80-285, 81-285, 82-285, 83-285, 84-285,85-285, 86-285, 87-285, 88-285, 89-285, 90-285, 91-285, 92-285, 93-285,94-285, 95-285, 96-285, 97-285, 98-285, 99-285, 100-285, 101-285,102-285, 103-285, 104-285, 105-285, 106-285, 107-285, 108-285, 109-285,110-285, 111-285, 112-285, 113-285, 114-285, 115-285, 116-285, 117-285,118-285, 119-285, 120-285, 121-285, 122-285, 123-285, 124-285, 125-285,126-285, 127-285, 128-285, 129-285, 130-285, 131-285, 132-285, 133-285,134-285, 135-285, 136-285, 137-285, 138-285, 139-285, 140-285, 141-285,142-285, 143-285, 144-285, 145-285, 146-285, 147-285, 148-285, 149-285,150-285, 151-285, 152-285, 153-285, 154-285, 155-285, 156-285, 157-285,158-285, 159-285, 160-285, 161-285, 162-285, 163-285, 164-285, 165-285,166-285, 167-285, 168-285, 169-285, 170-285, 171-285, 172-285, 173-285,174-285, 175-285, 176-285, 177-285, 178-285, 179-285, 180-285, 181-285,182-285, 183-285, 184-285, 185-285, 186-285, 187-285, 188-285, 189-285,and 190-285 of SEQ ID NO:3228. The present invention is also directed toantibodies that bind B Lymphocyte Stimulator polypeptides comprising, oralternatively, consisting of, a contiguous sequence of amino acidresidues at least 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99%identical to the amino acid sequence of B Lymphocyte Stimulatorpolypeptides described above.

Furthermore, since the predicted extracellular domain of the BLymphocyte Stimulator polypeptides of the invention may itself elicitbiological activity, deletions of N- and C-terminal amino acid residuesfrom the predicted extracellular region of the polypeptide (spanningpositions Gln-73 to Leu-285 of SEQ ID NO:3228) may retain somebiological activity such as, for example, ligand binding, stimulation oflymphocyte (e.g., B cell) proliferation, differentiation, and/oractivation, and modulation of cell replication or modulation of targetcell activities. However, even if deletion of one or more amino acidsfrom the N-terminus of the predicted extracellular domain of a BLymphocyte Stimulator polypeptide results in modification or loss of oneor more biological functions of the polypeptide, other functionalactivities may still be retained. Thus, the ability of the shortenedpolypeptides to induce and/or bind to antibodies which recognize thecomplete or mature or extracellular domains of the polypeptidesgenerally will be retained when less than the majority of the residuesof the complete or mature or extracellular domains of the polypeptidesare removed from the N-terminus. Whether a particular polypeptidelacking N-terminal residues of a complete polypeptide retains suchimmunologic activities can readily be determined by routine methodsdescribed herein and otherwise known in the art.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues deleted from the amino terminusof the amino acid sequence of B Lymphocyte Stimulator shown in SEQ IDNO:3228, up to the glycine residue at position number 280. Inparticular, the present invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, the amino acidsequence of residues n²-285 of SEQ ID NO:3228, where n² is an integer inthe range of the amino acid position of amino acid residues 73-280 inSEQ ID NO:3228, and 73 is the position of the first residue from theN-terminus of the predicted extracellular domain of the B LymphocyteStimulator polypeptide (disclosed in SEQ ID NO:3228). More inparticular, in certain embodiments, the invention provides antibodiesthat bind polypeptides comprising, or alternatively consisting of, anamino acid sequence selected from the group consisting of residues ofQ-73 to L-285; G-74 to L-285; D-75 to L-285; L-76 to L-285; A-77 toL-285; S-78 to L-285; L-79 to L-285; R-80 to L-285; A-81 to L-285; E-82to L-285; L-83 to L-285; Q-84 to L-285; G-85 to L-285; H-86 to L-285;H-87 to L-285; A-88 to L-285; E-89 to L-285; K-90 to L-285; L-91 toL-285; P-92 to L-285; A-93 to L-285; G-94 to L-285; A-95 to L-285; G-96to L-285; A-97 to L-285; P-98 to L-285; K-99 to L-285; A-100 to L-285;G-101 to L-285; L-102 to L-285; E-103 to L-285; E-104 to L-285; A-105 toL-285; P-106 to L-285; A-107 to L-285; V-108 to L-285; T-109 to L-285;A-110 to L-285; G-111 to L-285; L-112 to L-285; K-113 to L-285; I-114 toL-285; F-115 to L-285; E-116 to L-285; P-117 to L-285; P-118 to L-285;A-119 to L-285; P-120 to L-285; G-121 to L-285; E-122 to L-285; G-123 toL-285; N-124 to L-285; S-125 to L-285; S-126 to L-285; Q-127 to L-285;N-128 to L-285; S-129 to L-285; R-130 to L-285; N-131 to L-285; K-132 toL-285; R-133 to L-285; A-134 to L-285; V-135 to L-285; Q-136 to L-285;G-137 to L-285; P-138 to L-285; E-139 to L-285; E-140 to L-285; T-141 toL-285; V-142 to L-285; T-143 to L-285; Q-144 to L-285; D-145 to L-285;C-146 to L-285; L-147 to L-285; Q-148 to L-285; L-149 to L-285; I-150 toL-285; A-151 to L-285; D-152 to L-285; S-153 to L-285; E-154 to L-285;T-155 to L-285; P-156 to L-285; T-157 to L-285; I-158 to L-285; Q-159 toL-285; K-160 to L-285; G-161 to L-285; S-162 to L-285; Y-163 to L-285;T-164 to L-285; F-165 to L-285; V-166 to L-285; P-167 to L-285; W-168 toL-285; L-169 to L-285; L-170 to L-285; S-171 to L-285; F-172 to L-285;K-173 to L-285; R-174 to L-285; G-175 to L-285; S-176 to L-285; A-177 toL-285; L-178 to L-285; E-179 to L-285; E-180 to L-285; K-181 to L-285;E-182 to L-285; N-183 to L-285; K-184 to L-285; I-185 to L-285; L-186 toL-285; V-187 to L-285; K-188 to L-285; E-189 to L-285; T-190 to L-285;G-191 to L-285; Y-192 to L-285; F-193 to L-285; F-194 to L-285; I-195 toL-285; Y-196 to L-285; G-197 to L-285; Q-198 to L-285; V-199 to L-285;L-200 to L-285; Y-201 to L-285; T-202 to L-285; D-203 to L-285; K-204 toL-285; T-205 to L-285; Y-206 to L-285; A-207 to L-285; M-208 to L-285;G-209 to L-285; H-210 to L-285; L-211 to L-285; I-212 to L-285; Q-213 toL-285; R-214 to L-285; K-215 to L-285; K-216 to L-285; V-217 to L-285;H-218 to L-285; V-219 to L-285; F-220 to L-285; G-221 to L-285; D-222 toL-285; E-223 to L-285; L-224 to L-285; S-225 to L-285; L-226 to L-285;V-227 to L-285; T-228 to L-285; L-229 to L-285; F-230 to L-285; R-231 toL-285; C-232 to L-285; I-233 to L-285; Q-234 to L-285; N-235 to L-285;M-236 to L-285; P-237 to L-285; E-238 to L-285; T-239 to L-285; L-240 toL-285; P-241 to L-285; N-242 to L-285; N-243 to L-285; S-244 to L-285;C-245 to L-285; Y-246 to L-285; S-247 to L-285; A-248 to L-285; G-249 toL-285; I-250 to L-285; A-251 to L-285; K-252 to L-285; L-253 to L-285;E-254 to L-285; E-255 to L-285; G-256 to L-285; D-257 to L-285; E-258 toL-285; L-259 to L-285; Q-260 to L-285; L-261 to L-285; A-262 to L-285;1-263 to L-285; P-264 to L-285; R-265 to L-285; E-266 to L-285; N-267 toL-285; A-268 to L-285; Q-269 to L-285; I-270 to L-285; S-271 to L-285;L-272 to L-285; D-273 to L-285; G-274 to L-285; D-275 to L-285; V-276 toL-285; T-277 to L-285; F-278 to L-285; F-279 to L-285; and G-280 toL-285 of SEQ ID NO:3228. The present invention is also directed toantibodies that bind B Lymphocyte Stimulator polypeptides comprising, oralternatively, consisting of, a contiguous sequence of amino acidresidues at least 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99%identical to the amino acid sequence of B Lymphocyte Stimulatorpolypeptides described above.

Highly preferred embodiments of the invention are directed to antibodiesthat bind polypeptides comprising, or alternatively consisting of, apolypeptide having an amino acid sequence least 80%, 85%, 90% identicaland more preferably at least 95%, 96%, 97%, 98%, 99% or 100% identicalto B Lymphocyte Stimulator polypeptide having the amino acid sequence atpositions 134-285 of SEQ ID NO:3228.

Preferred embodiments of the invention are directed to antibodies thatbind polypeptides comprising, or alternatively consisting of, apolypeptide having an amino acid sequence at least 90% identical to a BLymphocyte Stimulator polypeptide having the amino acid sequence atpositions 134-285 of SEQ ID NO:3228. More preferred embodiments of theinvention are directed to antibodies that bind polypeptides comprising,or alternatively consisting of, a polypeptide having an amino acidsequence at least 95% identical to a B Lymphocyte Stimulator polypeptidehaving the amino acid sequence at positions 134-285 of SEQ ID NO:3228.More preferred embodiments of the invention are directed to antibodiesthat bind polypeptides comprising, or alternatively consisting of, apolypeptide having an amino acid sequence at least 96% identical to a BLymphocyte Stimulator polypeptide having the amino acid sequence atpositions 134-285 of SEQ ID NO:3228.

Additionally, more preferred embodiments of the invention are directedto antibodies that bind polypeptides comprising, or alternativelyconsisting of, a polypeptide having an amino acid sequence at least 97%to a B Lymphocyte Stimulator polypeptide having the amino acid sequenceat positions 134-285 of SEQ ID NO:3228. Additionally, more preferredembodiments of the invention are directed to antibodies that bindpolypeptides comprising, or alternatively consisting of, a polypeptidehaving an amino acid sequence at least 98% to a B Lymphocyte Stimulatorpolypeptide having the amino acid sequence at positions 134-285 of SEQID NO:3228. Additionally, more preferred embodiments of the inventionare directed to antibodies that bind polypeptides comprising, oralternatively consisting of, a polypeptide having an amino acid sequenceat least 99% identical to B Lymphocyte Stimulator polypeptide having theamino acid sequence at positions 134-285 of SEQ ID NO:3228.

In specific embodiments, antibodies of the present invention bindpolypeptides comprising, or alternatively consisting of, one of thefollowing N-terminally deleted polypeptide fragments of B LymphocyteStimulator amino acid residues Ala-71 through Leu-285, amino acidresidues Ala-81 through Leu-285, amino acid residues Leu-112 throughLeu-285, amino acid residues Ala-134 through Leu-285, amino acidresidues Leu-147 through Leu-285, and amino acid residues Gly-161through Leu-285 of SEQ ID NO:3228.

Similarly, many examples of biologically functional C-terminal deletionpolypeptides are known. For instance, Interferon gamma shows up to tentimes higher activities by deleting 8-10 amino acid residues from thecarboxy terminus of the protein (Dobeli et al., J. Biotechnology7:199-216 (1988). Since the present protein is a member of the TNFpolypeptide family, deletions of C-terminal amino acids up to theleucine residue at position 284 are expected to retain most if not allbiological activity such as, for example, ligand binding, the ability tostimulate lymphocyte (e.g., B cell) proliferation, differentiation,and/or activation, and modulation of cell replication. Polypeptideshaving deletions of up to about 10 additional C-terminal residues (i.e.,up to the glycine residue at position 274) also may retain some activitysuch as receptor binding, although such polypeptides would lack aportion of the conserved TNF domain which extends to about Leu-284 ofSEQ ID NO:3228. However, even if deletion of one or more amino acidsfrom the C-terminus of a protein results in modification or loss of oneor more biological functions of the protein, other functional activitiesmay still be retained. Thus, the ability of the shortened protein toinduce and/or bind to antibodies which recognize the complete or matureprotein generally will be retained when less than the majority of theresidues of the complete or mature protein are removed from theC-terminus. Whether a particular polypeptide lacking C-terminal residuesof a complete protein retains such immunologic activities can readily bedetermined by routine methods described herein and otherwise known inthe art.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues deleted from the carboxyterminus of the amino acid sequence of the B Lymphocyte Stimulatorpolypeptide of SEQ ID NO:3228, up to the glycine residue at position 274(Gly-274). In particular, the present invention provides antibodies thatbind polypeptides comprising, or alternatively consisting of, the aminoacid sequence of residues 1-m¹ of the amino acid sequence in SEQ IDNO:3228, where m¹ is any integer in the range of the amino acid positionof amino acid residues 274-284 in SEQ ID NO:3228. More in particular,the invention provides antibodies that bind B Lymphocyte Stimulatorpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues 1-274, 1-275,1-276, 1-277, 1-278, 1-279, 1-280, 1-281, 1-282, 1-283 and 1-284 of SEQID NO:3228. The present invention is also directed to antibodies thatbind B Lymphocyte Stimulator polypeptides comprising, or alternatively,consisting of, a contiguous sequence of amino acid residues at least80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99% identical to the aminoacid sequence of B Lymphocyte Stimulator polypeptides described above.

Also provided are antibodies that bind B Lymphocyte Stimulatorpolypeptides comprising, or alternatively consisting of, B LymphocyteStimulator polypeptides with one or more amino acids deleted from boththe amino and the carboxyl termini, which may be described generally ashaving residues n¹-m¹ of SEQ ID NO:3228, where n¹ and m¹ are integers asdefined above. Also included are antibodies that bind a polypeptidecomprising, or alternatively consisting of, a portion of the complete BLymphocyte Stimulator amino acid sequence encoded by the deposited cDNAclone contained in ATCC™ Accession No. 97768 where this portion excludesfrom 1 to 190 amino acids from the amino terminus or from 1 to 11 aminoacids from the C-terminus of the complete amino acid sequence (or anycombination of these N-terminal and C-terminal deletions) encoded by thecDNA clone in the deposited plasmid.

Similarly, deletions of C-terminal amino acid residues of the predictedextracellular domain of B Lymphocyte Stimulator up to the leucineresidue at position 79 of SEQ ID NO:3228 may retain some biologicalactivity, such as, for example, ligand binding, stimulation oflymphocyte (e.g., B cell) proliferation, differentiation, and/oractivation, and modulation of cell replication or modulation of targetcell activities. Polypeptides having further C-terminal deletionsincluding Leu-79 of SEQ ID NO:3228 would not be expected to retainbiological activities.

However, even if deletion of one or more amino acids from the C-terminusof a polypeptide results in modification or loss of one or morebiological functions of the polypeptide, other functional activities maystill be retained. Thus, the ability of the shortened polypeptide toinduce and/or bind to antibodies which recognize the complete, mature orextracellular forms of the polypeptide generally will be retained whenless than the majority of the residues of the complete, mature orextracellular forms of the polypeptide are removed from the C-terminus.Whether a particular polypeptide lacking C-terminal residues of thepredicted extracellular domain retains such immunologic activities canreadily be determined by routine methods described herein and otherwiseknown in the art.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues deleted from the carboxyterminus of the amino acid sequence of the predicted extracellulardomain of B Lymphocyte Stimulator polypeptide shown in SEQ ID NO:3228,up to the leucine residue at position 79 of SEQ ID NO:3228. Inparticular, the present invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, the amino acidsequence of residues 73-m² of the amino acid sequence in SEQ ID NO:3228,where m² is any integer in the range of the amino acid position of aminoacid residues 79 to 285 in the amino acid sequence in SEQ ID NO:3228,and residue 78 is the position of the first residue at the C-terminus ofthe predicted extracellular domain of the B Lymphocyte Stimulatorpolypeptide (disclosed in SEQ ID NO:3228). More in particular, incertain embodiments, the invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues Q-73 to Leu-285;Q-73 to L-284; Q-73 to K-283; Q-73 to L-282; Q-73 to A-281; Q-73 toG-280; Q-73 to F-279; Q-73 to F-278; Q-73 to T-277; Q-73 to V-276; Q-73to D-275; Q-73 to G-274; Q-73 to D-273; Q-73 to L-272; Q-73 to S-271;Q-73 to I-270; Q-73 to Q-269; Q-73 to A-268; Q-73 to N-267; Q-73 toE-266; Q-73 to R-265; Q-73 to P-264; Q-73 to I-263; Q-73 to A-262; Q-73to L-261; Q-73 to Q-260; Q-73 to L-259; Q-73 to E-258; Q-73 to D-257;Q-73 to G-256; Q-73 to E-255; Q-73 to E-254; Q-73 to L-253; Q-73 toK-252; Q-73 to A-251; Q-73 to I-250; Q-73 to G-249; Q-73 to A-248; Q-73to S-247; Q-73 to Y-246; Q-73 to C-245; Q-73 to S-244; Q-73 to N-243;Q-73 to N-242; Q-73 to P-241; Q-73 to L-240; Q-73 to T-239; Q-73 toE-238; Q-73 to P-237; Q-73 to M-236; Q-73 to N-235; Q-73 to Q-234; Q-73to I-233; Q-73 to C-232; Q-73 to R-231; Q-73 to F-230; Q-73 to L-229;Q-73 to T-228; Q-73 to V-227; Q-73 to L-226; Q-73 to S-225; Q-73 toL-224; Q-73 to E-223; Q-73 to D-222; Q-73 to G-221; Q-73 to F-220; Q-73to V-219; Q-73 to H-218; Q-73 to V-217; Q-73 to K-216; Q-73 to K-215;Q-73 to R-214; Q-73 to Q-213; Q-73 to 1-212; Q-73 to L-211; Q-73 toH-210; Q-73 to G-209; Q-73 to M-208; Q-73 to A-207; Q-73 to Y-206; Q-73to T-205; Q-73 to K-204; Q-73 to D-203; Q-73 to T-202; Q-73 to Y-201;Q-73 to L-200; Q-73 to V-199; Q-73 to Q-198; Q-73 to G-197; Q-73 toY-196; Q-73 to I-195; Q-73 to F-194; Q-73 to F-193; Q-73 to Y-192; Q-73to G-191; Q-73 to T-190; Q-73 to E-189; Q-73 to K-188; Q-73 to V-187;Q-73 to L-186; Q-73 to I-185; Q-73 to K-184; Q-73 to N-183; Q-73 toE-182; Q-73 to K-181; Q-73 to E-180; Q-73 to E-179; Q-73 to L-178; Q-73to A-177; Q-73 to S-176; Q-73 to G-175; Q-73 to R-174; Q-73 to K-173;Q-73 to F-172; Q-73 to S-171; Q-73 to L-170; Q-73 to L-169; Q-73 toW-168; Q-73 to P-167; Q-73 to V-166; Q-73 to F-165; Q-73 to T-164; Q-73to Y-163; Q-73 to S-162; Q-73 to G-161; Q-73 to K-160; Q-73 to Q-159;Q-73 to I-158; Q-73 to T-157; Q-73 to P-156; Q-73 to T-155; Q-73 toE-154; Q-73 to S-153; Q-73 to D-152; Q-73 to A-151; Q-73 to I-150; Q-73to L-149; Q-73 to Q-148; Q-73 to L-147; Q-73 to C-146; Q-73 to D-145;Q-73 to Q-144; Q-73 to T-143; Q-73 to V-142; Q-73 to T-141; Q-73 toE-140; Q-73 to E-139; Q-73 to P-138; Q-73 to G-137; Q-73 to Q-136; Q-73to V-135; Q-73 to A-134; Q-73 to R-133; Q-73 to K-132; Q-73 to N-131;Q-73 to R-130; Q-73 to S-129; Q-73 to N-128; Q-73 to Q-127; Q-73 toS-126; Q-73 to S-125; Q-73 to N-124; Q-73 to G-123; Q-73 to E-122; Q-73to G-121; Q-73 to P-120; Q-73 to A-119; Q-73 to P-118; Q-73 to P-117;Q-73 to E-116; Q-73 to F-115; Q-73 to I-114; Q-73 to K-113; Q-73 toL-112; Q-73 to G-111; Q-73 to A-110; Q-73 to T-109; Q-73 to V-108; Q-73to A-107; Q-73 to P-106; Q-73 to A-105; Q-73 to E-104; Q-73 to E-103;Q-73 to L-102; Q-73 to G-101; Q-73 to A-100; Q-73 to K-99; Q-73 to P-98;Q-73 to A-97; Q-73 to G-96; Q-73 to A-95; Q-73 to G-94; Q-73 to A-93;Q-73 to P-92; Q-73 to L-91; Q-73 to K-90; Q-73 to E-89; Q-73 to A-88;Q-73 to H-87; Q-73 to H-86; Q-73 to G-85; Q-73 to Q-84; Q-73 to L-83;Q-73 to E-82; Q-73 to A-81; Q-73 to R-80; and Q-73 to L-79 of SEQ IDNO:3228. The present invention is also directed to antibodies that bindB Lymphocyte Stimulator polypeptides comprising, or alternatively,consisting of, a contiguous sequence of amino acid residues at least80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99% identical to the aminoacid sequence of B Lymphocyte Stimulator polypeptides described above.

The invention also provides antibodies that bind polypeptides having oneor more amino acids deleted from both the amino and the carboxyl terminiof the predicted extracellular domain of B Lymphocyte Stimulator, whichmay be described generally as having residues n²-m² of SEQ ID NO:3228where n² and m² are integers as defined above.

In another embodiment, antibodies of the present invention bindpolypeptides consisting of a portion of the extracellular domain of theB Lymphocyte Stimulator amino acid sequence encoded by the cDNA plasmidcontained in the deposit having ATCC™ accession no. 97768, where thisportion excludes from 1 to about 206 amino acids from the amino terminusof the extracellular domain of the amino acid sequence encoded by thecDNA plasmid contained in the deposit having ATCC™ accession no. 97768,or from 1 to about 206 amino acids from the carboxy terminus of theextracellular domain of the amino acid sequence encoded by the cDNAplasmid contained in the deposit having ATCC™ accession no. 97768, orany combination of the above amino terminal and carboxy terminaldeletions, of the entire extracellular domain of the amino acid sequenceencoded by the cDNA plasmid contained in the deposit having ATCC™accession no. 97768.

As mentioned above, even if deletion of one or more amino acids from theN-terminus of a polypeptide results in modification or loss of one ormore functional activities (e.g., biological activity) of thepolypeptide, other functions or biological activities may still beretained. Thus, the ability of a shortened B Lymphocyte Stimulatormutein to induce and/or bind to antibodies which recognize thefull-length or mature forms or the extracellular domain of thepolypeptide generally will be retained when less than the majority ofthe residues of the full-length or mature or extracellular domain of thepolypeptide are removed from the N-terminus. Whether a particularpolypeptide lacking N-terminal residues of a complete polypeptideretains such immunologic activities can readily be determined by routinemethods described herein and otherwise known in the art. It is notunlikely that a B Lymphocyte Stimulator mutein with a large number ofdeleted N-terminal amino acid residues may retain some functional (e.g.,biological or immunogenic) activities. In fact, peptides composed of asfew as six B Lymphocyte Stimulator amino acid residues may often evokean immune response.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues deleted from the amino terminusof the predicted full-length amino acid sequence of the B LymphocyteStimulator shown in SEQ ID NO:3228, up to the glycine residue atposition number 280 of the sequence shown SEQ ID NO:3228 andpolynucleotides encoding such polypeptides. In particular, the presentinvention provides antibodies that bind polypeptides comprising theamino acid sequence of residues n³-285 of the sequence shown in SEQ IDNO:3228, where n³ is an integer in the range of the amino acid positionof amino acid residues 1 to 280 of the amino acid sequence in SEQ IDNO:3228.

More in particular, the invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues of D-2 to L-285;D-3 to L-285; S-4 to L-285; T-5 to L-285; E-6 to L-285; R-7 to L-285;E-8 to L-285; Q-9 to L-285; S-10 to L-285; R-11 to L-285; L-12 to L-285;T-13 to L-285; S-14 to L-285; C-15 to L-285; L-16 to L-285; K-17 toL-285; K-18 to L-285; R-19 to L-285; E-20 to L-285; E-21 to L-285; M-22to L-285; K-23 to L-285; L-24 to L-285; K-25 to L-285; E-26 to L-285;C-27 to L-285; V-28 to L-285; S-29 to L-285; 1-30 to L-285; L-31 toL-285; P-32 to L-285; R-33 to L-285; K-34 to L-285; E-35 to L-285; S-36to L-285; P-37 to L-285; S-38 to L-285; V-39 to L-285; R-40 to L-285;S-41 to L-285; S-42 to L-285; K-43 to L-285; D-44 to L-285; G-45 toL-285; K-46 to L-285; L-47 to L-285; L-48 to L-285; A-49 to L-285; A-50to L-285; T-51 to L-285; L-52 to L-285; L-53 to L-285; L-54 to L-285;A-55 to L-285; L-56 to L-285; L-57 to L-285; S-58 to L-285; C-59 toL-285; C-60 to L-285; L-61 to L-285; T-62 to L-285; V-63 to L-285; V-64to L-285; S-65 to L-285; F-66 to L-285; Y-67 to L-285; Q-68 to L-285;V-69 to L-285; A-70 to L-285; A-71 to L-285; L-72 to L-285; Q-73 toL-285; G-74 to L-285; D-75 to L-285; L-76 to L-285; A-77 to L-285; S-78to L-285; L-79 to L-285; R-80 to L-285; A-81 to L-285; E-82 to L-285;L-83 to L-285; Q-84 to L-285; G-85 to L-285; H-86 to L-285; H-87 toL-285; A-88 to L-285; E-89 to L-285; K-90 to L-285; L-91 to L-285; P-92to L-285; A-93 to L-285; G-94 to L-285; A-95 to L-285; G-96 to L-285;A-97 to L-285; P-98 to L-285; K-99 to L-285; A-100 to L-285; G-101 toL-285; L-102 to L-285; E-103 to L-285; E-104 to L-285; A-105 to L-285;P-106 to L-285; A-107 to L-285; V-108 to L-285; T-109 to L-285; A-110 toL-285; G-111 to L-285; L-112 to L-285; K-113 to L-285; I-114 to L-285;F-115 to L-285; E-116 to L-285; P-117 to L-285; P-118 to L-285; A-119 toL-285; P-120 to L-285; G-121 to L-285; E-122 to L-285; G-123 to L-285;N-124 to L-285; S-125 to L-285; S-126 to L-285; Q-127 to L-285; N-128 toL-285; S-129 to L-285; R-130 to L-285; N-131 to L-285; K-132 to L-285;R-133 to L-285; A-134 to L-285; V-135 to L-285; Q-136 to L-285; G-137 toL-285; P-138 to L-285; E-139 to L-285; E-140 to L-285; T-141 to L-285;V-142 to L-285; T-143 to L-285; Q-144 to L-285; D-145 to L-285; C-146 toL-285; L-147 to L-285; Q-148 to L-285; L-149 to L-285; I-150 to L-285;A-151 to L-285; D-152 to L-285; S-153 to L-285; E-154 to L-285; T-155 toL-285; P-156 to L-285; T-157 to L-285; I-158 to L-285; Q-159 to L-285;K-160 to L-285; G-161 to L-285; S-162 to L-285; Y-163 to L-285; T-164 toL-285; F-165 to L-285; V-166 to L-285; P-167 to L-285; W-168 to L-285;L-169 to L-285; L-170 to L-285; S-171 to L-285; F-172 to L-285; K-173 toL-285; R-174 to L-285; G-175 to L-285; S-176 to L-285; A-177 to L-285;L-178 to L-285; E-179 to L-285; E-180 to L-285; K-181 to L-285; E-182 toL-285; N-183 to L-285; K-184 to L-285; 1-185 to L-285; L-186 to L-285;V-187 to L-285; K-188 to L-285; E-189 to L-285; T-190 to L-285; G-191 toL-285; Y-192 to L-285; F-193 to L-285; F-194 to L-285; I-195 to L-285;Y-196 to L-285; G-197 to L-285; Q-198 to L-285; V-199 to L-285; L-200 toL-285; Y-201 to L-285; T-202 to L-285; D-203 to L-285; K-204 to L-285;T-205 to L-285; Y-206 to L-285; A-207 to L-285; M-208 to L-285; G-209 toL-285; H-210 to L-285; L-211 to L-285; I-212 to L-285; Q-213 to L-285;R-214 to L-285; K-215 to L-285; K-216 to L-285; V-217 to L-285; H-218 toL-285; V-219 to L-285; F-220 to L-285; G-221 to L-285; D-222 to L-285;E-223 to L-285; L-224 to L-285; S-225 to L-285; L-226 to L-285; V-227 toL-285; T-228 to L-285; L-229 to L-285; F-230 to L-285; R-231 to L-285;C-232 to L-285; I-233 to L-285; Q-234 to L-285; N-235 to L-285; M-236 toL-285; P-237 to L-285; E-238 to L-285; T-239 to L-285; L-240 to L-285;P-241 to L-285; N-242 to L-285; N-243 to L-285; S-244 to L-285; C-245 toL-285; Y-246 to L-285; S-247 to L-285; A-248 to L-285; G-249 to L-285;I-250 to L-285; A-251 to L-285; K-252 to L-285; L-253 to L-285; E-254 toL-285; E-255 to L-285; G-256 to L-285; D-257 to L-285; E-258 to L-285;L-259 to L-285; Q-260 to L-285; L-261 to L-285; A-262 to L-285; I-263 toL-285; P-264 to L-285; R-265 to L-285; E-266 to L-285; N-267 to L-285;A-268 to L-285; Q-269 to L-285; I-270 to L-285; S-271 to L-285; L-272 toL-285; D-273 to L-285; G-274 to L-285; D-275 to L-285; V-276 to L-285;T-277 to L-285; F-278 to L-285; F-279 to L-285; and G-280 to L-285 ofSEQ ID NO:3228. The present invention is also directed to antibodiesthat bind B Lymphocyte Stimulator polypeptides comprising, oralternatively, consisting of, a contiguous sequence of amino acidresidues at least 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99%identical to the amino acid sequence of B Lymphocyte Stimulatorpolypeptides described above.

Also as mentioned above, even if deletion of one or more amino acidsfrom the C-terminus of a protein results in modification or loss of oneor more functional activities (e.g., biological activity) of theprotein, other functional activities may still be retained. Thus, theability of a shortened B Lymphocyte Stimulator mutein to induce and/orbind to antibodies which recognize the complete or mature form or theextracellular domain of the polypeptide generally will be retained whenless than the majority of the residues of the complete or mature form orthe extracellular domain of the polypeptide are removed from theC-terminus. Whether a particular polypeptide lacking C-terminal residuesof a complete polypeptide retains such immunologic activities canreadily be determined by routine methods described herein and otherwiseknown in the art. It is not unlikely that a B Lymphocyte Stimulatormutein with a large number of deleted C-terminal amino acid residues mayretain some functional (e.g., biological or immunogenic) activities. Infact, peptides composed of as few as six B Lymphocyte Stimulator aminoacid residues may often evoke an immune response.

Accordingly, the present invention further provides in anotherembodiment, antibodies that bind polypeptides having one or moreresidues deleted from the carboxy terminus of the amino acid sequence ofthe B Lymphocyte Stimulator shown in SEQ ID NO:3228, up to the glutamicacid residue at position number 6, and polynucleotides encoding suchpolypeptides. In particular, the present invention provides antibodiesthat bind polypeptides comprising the amino acid sequence of residues1-m³ of SEQ ID NO:3228, where m³ is an integer in the range of the aminoacid position of amino acid residues 6-284 of the amino acid sequence inSEQ ID NO:3228.

More in particular, the invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues M-1 to L-284;M-1 to K-283; M-1 to L-282; M-1 to A-281; M-1 to G-280; M-1 to F-279;M-1 to F-278; M-1 to T-277; M-1 to V-276; M-1 to D-275; M-1 to G-274;M-1 to D-273; M-1 to L-272; M-1 to S-271; M-1 to I-270; M-1 to Q-269;M-1 to A-268; M-1 to N-267; M-1 to E-266; M-1 to R-265; M-1 to P-264;M-1 to I-263; M-1 to A-262; M-1 to L-261; M-1 to Q-260; M-1 to L-259;M-1 to E-258; M-1 to D-257; M-1 to G-256; M-1 to E-255; M-1 to E-254;M-1 to L-253; M-1 to K-252; M-1 to A-251; M-1 to I-250; M-1 to G-249;M-1 to A-248; M-1 to S-247; M-1 to Y-246; M-1 to C-245; M-1 to S-244;M-1 to N-243; M-1 to N-242; M-1 to P-241; M-1 to L-240; M-1 to T-239;M-1 to E-238; M-1 to P-237; M-1 to M-236; M-1 to N-235; M-1 to Q-234;M-1 to I-233; M-1 to C-232; M-1 to R-231; M-1 to F-230; M-1 to L-229;M-1 to T-228; M-1 to V-227; M-1 to L-226; M-1 to S-225; M-1 to L-224;M-1 to E-223; M-1 to D-222; M-1 to G-221; M-1 to F-220; M-1 to V-219;M-1 to H-218; M-1 to V-217; M-1 to K-216; M-1 to K-215; M-1 to R-214;M-1 to Q-213; M-1 to I-212; M-1 to L-211; M-1 to H-210; M-1 to G-209;M-1 to M-208; M-1 to A-207; M-1 to Y-206; M-1 to T-205; M-1 to K-204;M-1 to D-203; M-1 to T-202; M-1 to Y-201; M-1 to L-200; M-1 to V-199;M-1 to Q-198; M-1 to G-197; M-1 to Y-196; M-1 to I-195; M-1 to F-194;M-1 to F-193; M-1 to Y-192; M-1 to G-191; M-1 to T-190; M-1 to E-189;M-1 to K-188; M-1 to V-187; M-1 to L-186; M-1 to I-185; M-1 to K-184;M-1 to N-183; M-1 to E-182; M-1 to K-181; M-1 to E-180; M-1 to E-179;M-1 to L-178; M-1 to A-177; M-1 to S-176; M-1 to G-175; M-1 to R-174;M-1 to K-173; M-1 to F-172; M-1 to S-171; M-1 to L-170; M-1 to L-169;M-1 to W-168; M-1 to P-167; M-1 to V-166; M-1 to F-165; M-1 to T-164;M-1 to Y-163; M-1 to S-162; M-1 to G-161; M-1 to K-160; M-1 to Q-159;M-1 to I-158; M-1 to T-157; M-1 to P-156; M-1 to T-155; M-1 to E-154;M-1 to S-153; M-1 to D-152; M-1 to A-151; M-1 to I-150; M-1 to L-149;M-1 to Q-148; M-1 to L-147; M-1 to C-146; M-1 to D-145; M-1 to Q-144;M-1 to T-143; M-1 to V-142; M-1 to T-141; M-1 to E-140; M-1 to E-139;M-1 to P-138; M-1 to G-137; M-1 to Q-136; M-1 to V-135; M-1 to A-134;M-1 to R-133; M-1 to K-132; M-1 to N-131; M-1 to R-130; M-1 to S-129;M-1 to N-128; M-1 to Q-127; M-1 to S-126; M-1 to S-125; M-1 to N-124;M-1 to G-123; M-1 to E-122; M-1 to G-121; M-1 to P-120; M-1 to A-119;M-1 to P-118; M-1 to P-117; M-1 to E-116; M-1 to F-115; M-1 to I-114;M-1 to K-113; M-1 to L-112; M-1 to G-111; M-1 to A-110; M-1 to T-109;M-1 to V-108; M-1 to A-107; M-1 to P-106; M-1 to A-105; M-1 to E-104;M-1 to E-103; M-1 to L-102; M-1 to G-101; M-1 to A-100; M-1 to K-99; M-1to P-98; M-1 to A-97; M-1 to G-96; M-1 to A-95; M-1 to G-94; M-1 toA-93; M-1 to P-92; M-1 to L-91; M-1 to K-90; M-1 to E-89; M-1 to A-88;M-1 to H-87; M-1 to H-86; M-1 to G-85; M-1 to Q-84; M-1 to L-83; M-1 toE-82; M-1 to A-81; M-1 to R-80; M-1 to L-79; M-1 to S-78; M-1 to A-77;M-1 to L-76; M-1 to D-75; M-1 to G-74; M-1 to Q-73; M-1 to L-72; M-1 toA-71; M-1 to A-70; M-1 to V-69; M-1 to Q-68; M-1 to Y-67; M-1 to F-66;M-1 to S-65; M-1 to V-64; M-1 to V-63; M-1 to T-62; M-1 to L-61; M-1 toC-60; M-1 to C-59; M-1 to S-58; M-1 to L-57; M-1 to L-56; M-1 to A-55;M-1 to L-54; M-1 to L-53; M-1 to L-52; M-1 to T-51; M-1 to A-50; M-1 toA-49; M-1 to L-48; M-1 to L-47; M-1 to K-46; M-1 to G-45; M-1 to D-44;M-1 to K-43; M-1 to S-42; M-1 to S-41; M-1 to R-40; M-1 to V-39; M-1 toS-38; M-1 to P-37; M-1 to S-36; M-1 to E-35; M-1 to K-34; M-1 to R-33;M-1 to P-32; M-1 to L-31; M-1 to I-30; M-1 to S-29; M-1 to V-28; M-1 toC-27; M-1 to E-26; M-1 to K-25; M-1 to L-24; M-1 to K-23; M-1 to M-22;M-1 to E-21; M-1 to E-20; M-1 to R-19; M-1 to K-18; M-1 to K-17; M-1 toL-16; M-1 to C-15; M-1 to S-14; M-1 to T-13; M-1 to L-12; M-1 to R-11;M-1 to S-10; M-1 to Q-9; M-1 to E-8; M-1 to R-7; and M-1 to E-6 of SEQID NO:3228. The present invention is also directed to antibodies thatbind B Lymphocyte Stimulator polypeptides comprising, or alternatively,consisting of, a contiguous sequence of amino acid residues at least80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99% identical to the aminoacid sequence of B Lymphocyte Stimulator polypeptides described above.

The invention also provides antibodies that bind polypeptides having oneor more amino acids deleted from both the amino and the carboxyl terminiof a B Lymphocyte Stimulator polypeptide, which may be describedgenerally as having residues n³-m³ of SEQ ID NO:3228, where n³ and m³are integers as defined above.

Furthermore, since the predicted extracellular domain of the BLymphocyte Stimulator polypeptide of SEQ ID NO:3229 may itself elicitfunctional activity (e.g., biological activity), deletions of N- andC-terminal amino acid residues from the predicted extracellular regionof the polypeptide at positions Gln-73 to Leu-266 of SEQ ID NO:3229 mayretain some functional activity, such as, for example, ligand binding,to stimulation of lymphocyte (e.g., B cell) proliferation,differentiation, and/or activation, modulation of cell replication,modulation of target cell activities and/or immunogenicity. However,even if deletion of one or more amino acids from the N-terminus of thepredicted extracellular domain of a B Lymphocyte Stimulator polypeptideresults in modification or loss of one or more functional activities ofthe polypeptide, other functional activities may still be retained.Thus, the ability of the shortened polypeptides to induce and/or bind toantibodies which recognize the complete or mature or extracellulardomains of the polypeptides generally will be retained when less thanthe majority of the residues of the complete or mature or extracellulardomains of the polypeptides are removed from the N-terminus. Whether aparticular polypeptide lacking N-terminal residues of a completepolypeptide retains such immunologic activities can readily bedetermined by routine methods described herein and otherwise known inthe art.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues deleted from the amino terminusof the amino acid sequence of B Lymphocyte Stimulator shown in SEQ IDNO:3229, up to the glycine residue at position number 261. Inparticular, the present invention provides antibodies that bindpolypeptides comprising the amino acid sequence of residues n⁴-266 ofSEQ ID NO:3229, where n⁴ is an integer in the range of the amino acidposition of amino acid residues 73-261 of the amino acid sequence in SEQID NO:3229, and 261 is the position of the first residue from theN-terminus of the predicted extracellular domain B Lymphocyte Stimulatorpolypeptide (shown in SEQ ID NO:3229).

More in particular, in certain embodiments, the invention providesantibodies that bind polypeptides comprising, or alternativelyconsisting of, an amino acid sequence selected from the group consistingof residues of Q-73 to L-266; G-74 to L-266; D-75 to L-266; L-76 toL-266; A-77 to L-266; S-78 to L-266; L-79 to L-266; R-80 to L-266; A-81to L-266; E-82 to L-266; L-83 to L-266; Q-84 to L-266; G-85 to L-266;H-86 to L-266; H-87 to L-266; A-88 to L-266; E-89 to L-266; K-90 toL-266; L-91 to L-266; P-92 to L-266; A-93 to L-266; G-94 to L-266; A-95to L-266; G-96 to L-266; A-97 to L-266; P-98 to L-266; K-99 to L-266;A-100 to L-266; G-101 to L-266; L-102 to L-266; E-103 to L-266; E-104 toL-266; A-105 to L-266; P-106 to L-266; A-107 to L-266; V-108 to L-266;T-109 to L-266; A-110 to L-266; G-111 to L-266; L-112 to L-266; K-113 toL-266; 1-114 to L-266; F-115 to L-266; E-116 to L-266; P-117 to L-266;P-118 to L-266; A-119 to L-266; P-120 to L-266; G-121 to L-266; E-122 toL-266; G-123 to L-266; N-124 to L-266; S-125 to L-266; S-126 to L-266;Q-127 to L-266; N-128 to L-266; S-129 to L-266; R-130 to L-266; N-131 toL-266; K-132 to L-266; R-133 to L-266; A-134 to L-266; V-135 to L-266;Q-136 to L-266; G-137 to L-266; P-138 to L-266; E-139 to L-266; E-140 toL-266; T-141 to L-266; G-142 to L-266; S-143 to L-266; Y-144 to L-266;T-145 to L-266; F-146 to L-266; V-147 to L-266; P-148 to L-266; W-149 toL-266; L-150 to L-266; L-151 to L-266; S-152 to L-266; F-153 to L-266;K-154 to L-266; R-155 to L-266; G-156 to L-266; S-157 to L-266; A-158 toL-266; L-159 to L-266; E-160 to L-266; E-161 to L-266; K-162 to L-266;E-163 to L-266; N-164 to L-266; K-165 to L-266; 1-166 to L-266; L-167 toL-266; V-168 to L-266; K-169 to L-266; E-170 to L-266; T-171 to L-266;G-172 to L-266; Y-173 to L-266; F-174 to L-266; F-175 to L-266; I-176 toL-266; Y-177 to L-266; G-178 to L-266; Q-179 to L-266; V-180 to L-266;L-181 to L-266; Y-182 to L-266; T-183 to L-266; D-184 to L-266; K-185 toL-266; T-186 to L-266; Y-187 to L-266; A-188 to L-266; M-189 to L-266;G-190 to L-266; H-191 to L-266; L-192 to L-266; I-193 to L-266; Q-194 toL-266; R-195 to L-266; K-196 to L-266; K-197 to L-266; V-198 to L-266;H-199 to L-266; V-200 to L-266; F-201 to L-266; G-202 to L-266; D-203 toL-266; E-204 to L-266; L-205 to L-266; S-206 to L-266; L-207 to L-266;V-208 to L-266; T-209 to L-266; L-210 to L-266; F-211 to L-266; R-212 toL-266; C-213 to L-266; I-214 to L-266; Q-215 to L-266; N-216 to L-266;M-217 to L-266; P-218 to L-266; E-219 to L-266; T-220 to L-266; L-221 toL-266; P-222 to L-266; N-223 to L-266; N-224 to L-266; S-225 to L-266;C-226 to L-266; Y-227 to L-266; S-228 to L-266; A-229 to L-266; G-230 toL-266; I-231 to L-266; A-232 to L-266; K-233 to L-266; L-234 to L-266;E-235 to L-266; E-236 to L-266; G-237 to L-266; D-238 to L-266; E-239 toL-266; L-240 to L-266; Q-241 to L-266; L-242 to L-266; A-243 to L-266;I-244 to L-266; P-245 to L-266; R-246 to L-266; E-247 to L-266; N-248 toL-266; A-249 to L-266; Q-250 to L-266; I-251 to L-266; S-252 to L-266;L-253 to L-266; D-254 to L-266; G-255 to L-266; D-256 to L-266; V-257 toL-266; T-258 to L-266; F-259 to L-266; F-260 to L-266; and G-261 toL-266 of SEQ ID NO:3229. The present invention is also directed toantibodies that bind B Lymphocyte Stimulator polypeptides comprising, oralternatively, consisting of, a contiguous sequence of amino acidresidues at least 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99%identical to the amino acid sequence of B Lymphocyte Stimulatorpolypeptides described above.

Similarly, deletions of C-terminal amino acid residues of the predictedextracellular domain of B Lymphocyte Stimulator up to the leucineresidue at position 79 of SEQ ID NO:3229 may retain some functionalactivity, such as, for example, ligand binding, the ability to stimulatelymphocyte (e.g., B cell) proliferation, differentiation, and/oractivation, modulation of cell replication, modulation of target cellactivities and/or immunogenicity. Polypeptides having further C-terminaldeletions including Leu-79 of SEQ ID NO:3229 would not be expected toretain biological activities.

However, even if deletion of one or more amino acids from the C-terminusof a polypeptide results in modification or loss of one or morefunctional activities (e.g., biological activity) of the polypeptide,other functional activities may still be retained. Thus, the ability ofthe shortened polypeptide to induce and/or bind to antibodies whichrecognize the complete, mature or extracellular forms of the polypeptidegenerally will be retained when less than the majority of the residuesof the complete, mature or extracellular forms of the polypeptide areremoved from the C-terminus. Whether a particular polypeptide lackingC-terminal residues of the predicted extracellular domain retains suchimmunologic activities can readily be determined by routine methodsdescribed herein and otherwise known in the art.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues from the carboxy terminus ofthe amino acid sequence of the predicted extracellular domain of BLymphocyte Stimulator shown in SEQ ID NO:3229, up to the leucine residueat position 79 of SEQ ID NO:3229. In particular, the present inventionprovides antibodies that bind polypeptides having the amino acidsequence of residues 73-m⁴ of the amino acid sequence in SEQ ID NO:3229,where m⁴ is any integer in the range of the amino acid position of aminoacid residues 79-265 of the amino acid sequence in SEQ ID NO:3229.

More in particular, in certain embodiments, the invention providesantibodies that bind polypeptides comprising, or alternativelyconsisting of, an amino acid sequence selected from the group consistingof residues Q-73 to L-265; Q-73 to K-264; Q-73 to L-263; Q-73 to A-262;Q-73 to G-261; Q-73 to F-260; Q-73 to F-259; Q-73 to T-258; Q-73 toV-257; Q-73 to D-256; Q-73 to G-255; Q-73 to D-254; Q-73 to L-253; Q-73to S-252; Q-73 to I-251; Q-73 to Q-250; Q-73 to A-249; Q-73 to N-248;Q-73 to E-247; Q-73 to R-246; Q-73 to P-245; Q-73 to I-244; Q-73 toA-243; Q-73 to L-242; Q-73 to Q-241; Q-73 to L-240; Q-73 to E-239; Q-73to D-238; Q-73 to G-237; Q-73 to E-236; Q-73 to E-235; Q-73 to L-234;Q-73 to K-233; Q-73 to A-232; Q-73 to I-231; Q-73 to G-230; Q-73 toA-229; Q-73 to S-228; Q-73 to Y-227; Q-73 to C-226; Q-73 to S-225; Q-73to N-224; Q-73 to N-223; Q-73 to P-222; Q-73 to L-221; Q-73 to T-220;Q-73 to E-219; Q-73 to P-218; Q-73 to M-217; Q-73 to N-216; Q-73 toQ-215; Q-73 to I-214; Q-73 to C-213; Q-73 to R-212; Q-73 to F-211; Q-73to L-210; Q-73 to T-209; Q-73 to V-208; Q-73 to L-207; Q-73 to S-206;Q-73 to L-205; Q-73 to E-204; Q-73 to D-203; Q-73 to G-202; Q-73 toF-201; Q-73 to V-200; Q-73 to H-199; Q-73 to V-198; Q-73 to K-197; Q-73to K-196; Q-73 to R-195; Q-73 to Q-194; Q-73 to I-193; Q-73 to L-192;Q-73 to H-191; Q-73 to G-190; Q-73 to Q-7389; Q-73 to A-188; Q-73 toY-187; Q-73 to T-186; Q-73 to K-185; Q-73 to D-184; Q-73 to T-183; Q-73to Y-182; Q-73 to L-181; Q-73 to V-180; Q-73 to Q-179; Q-73 to G-178;Q-73 to Y-177; Q-73 to I-176; Q-73 to F-175; Q-73 to F-174; Q-73 toY-173; Q-73 to G-172; Q-73 to T-171; Q-73 to E-170; Q-73 to K-169; Q-73to V-168; Q-73 to L-167; Q-73 to I-166; Q-73 to K-165; Q-73 to N-164;Q-73 to E-163; Q-73 to K-162; Q-73 to E-161; Q-73 to E-160; Q-73 toL-159; Q-73 to A-158; Q-73 to S-157; Q-73 to G-156; Q-73 to R-155; Q-73to K-154; Q-73 to F-153; Q-73 to S-152; Q-73 to L-151; Q-73 to L-150;Q-73 to W-149; Q-73 to P-148; Q-73 to V-147; Q-73 to F-146; Q-73 toT-145; Q-73 to Y-144; Q-73 to S-143; Q-73 to G-142; Q-73 to T-141; Q-73to E-140; Q-73 to E-139; Q-73 to P-138; Q-73 to G-137; Q-73 to Q-136;Q-73 to V-135; Q-73 to A-134; Q-73 to R-133; Q-73 to K-132; Q-73 toN-131; Q-73 to R-130; Q-73 to S-129; Q-73 to N-128; Q-73 to Q-127; Q-73to S-126; Q-73 to S-125; Q-73 to N-124; Q-73 to G-123; Q-73 to E-122;Q-73 to G-121; Q-73 to P-120; Q-73 to A-119; Q-73 to P-118; Q-73 toP-117; Q-73 to E-116; Q-73 to F-115; Q-73 to I-114; Q-73 to K-113; Q-73to L-112; Q-73 to G-111; Q-73 to A-110; Q-73 to T-109; Q-73 to V-108;Q-73 to A-107; Q-73 to P-106; Q-73 to A-105; Q-73 to E-104; Q-73 toE-103; Q-73 to L-102; Q-73 to G-101; Q-73 to A-100; Q-73 to K-99; Q-73to P-98; Q-73 to A-97; Q-73 to G-96; Q-73 to A-95; Q-73 to G-94; Q-73 toA-93; Q-73 to P-92; Q-73 to L-91; Q-73 to K-90; Q-73 to E-89; Q-73 toA-88; Q-73 to H-87; Q-73 to H-86; Q-73 to G-85; Q-73 to Q-84; Q-73 toL-83; Q-73 to E-82; Q-73 to A-81; Q-73 to R-80; Q-73 to L-79; and Q-73to S-78 of SEQ ID NO:3229. The present invention is also directed toantibodies that bind B Lymphocyte Stimulator polypeptides comprising, oralternatively, consisting of, a contiguous sequence of amino acidresidues at least 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99%identical to the amino acid sequence of B Lymphocyte Stimulatorpolypeptides described above.

The invention also provides polypeptides having one or more amino acidsdeleted from both the amino and the carboxyl termini of the predictedextracellular domain of B Lymphocyte Stimulator, which may be describedgenerally as having residues n⁴-m⁴ of SEQ ID NO:3229 where n⁴ and m⁴ areintegers as defined above.

In another embodiment, antibodies of the present invention bindpolypeptides consisting of a portion of the extracellular domain of theB Lymphocyte Stimulator amino acid sequence encoded by the cDNA clonecontained in the deposit having ATCC™ Accession No. 203518, where thisportion excludes from 1 to about 260 amino acids from the amino terminusof the extracellular domain of the amino acid sequence encoded by cDNAclone contained in the deposit having ATCC™ Accession No. 203518, orfrom 1 to about 187 amino acids from the carboxy terminus of theextracellular domain of the amino acid sequence encoded by cDNA clonecontained in the deposit having ATCC™ Accession No. 203518, or anycombination of the above amino terminal and carboxy terminal deletions,of the entire extracellular domain of the amino acid sequence encoded bythe cDNA clone contained in the deposit having ATCC™ Accession No.203518.

As mentioned above, even if deletion of one or more amino acids from theN-terminus of a polypeptide results in modification or loss of one ormore functional activities (e.g., biological activity) of thepolypeptide, other functional activities may still be retained. Thus,the ability of a shortened B Lymphocyte Stimulator polypeptide to induceand/or bind to antibodies which recognize the full-length or matureforms or the extracellular domain of the polypeptide generally will beretained when less than the majority of the residues of the full-lengthor mature or extracellular domain of the polypeptide are removed fromthe N-terminus. Whether a particular polypeptide lacking N-terminalresidues of a complete polypeptide retains such immunologic activitiescan readily be determined by routine methods described herein andotherwise known in the art. It is not unlikely that a B LymphocyteStimulator mutein with a large number of deleted N-terminal amino acidresidues may retain functional (e.g., immunogenic) activities. In fact,peptides composed of as few as six B Lymphocyte Stimulator amino acidresidues may often evoke an immune response.

Accordingly, the present invention further provides antibodies that bindpolypeptides having one or more residues deleted from the amino terminusof the predicted full-length amino acid sequence of the B LymphocyteStimulator polypeptide shown in SEQ ID NO:3229, up to the glycineresidue at position number 261 of the sequence shown SEQ ID NO:3229 andpolynucleotides encoding such polypeptides. In particular, the presentinvention provides antibodies that bind polypeptides comprising theamino acid sequence of residues n⁵-266 of the sequence shown in SEQ IDNO:3229, where n⁵ is an integer in the range of the amino acid positionof amino acid residues 1 to 261 of the amino acid sequence in SEQ IDNO:3229.

More in particular, the invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues of D-2 to L-266;D-3 to L-266; S-4 to L-266; T-5 to L-266; E-6 to L-266; R-7 to L-266;E-8 to L-266; Q-9 to L-266; S-10 to L-266; R-11 to L-266; L-12 to L-266;T-13 to L-266; S-14 to L-266; C-15 to L-266; L-16 to L-266; K-17 toL-266; K-18 to L-266; R-19 to L-266; E-20 to L-266; E-21 to L-266; M-22to L-266; K-23 to L-266; L-24 to L-266; K-25 to L-266; E-26 to L-266;C-27 to L-266; V-28 to L-266; S-29 to L-266; I-30 to L-266; L-31 toL-266; P-32 to L-266; R-33 to L-266; K-34 to L-266; E-35 to L-266; S-36to L-266; P-37 to L-266; S-38 to L-266; V-39 to L-266; R-40 to L-266;S-41 to L-266; S-42 to L-266; K-43 to L-266; D-44 to L-266; G-45 toL-266; K-46 to L-266; L-47 to L-266; L-48 to L-266; A-49 to L-266; A-50to L-266; T-51 to L-266; L-52 to L-266; L-53 to L-266; L-54 to L-266;A-55 to L-266; L-56 to L-266; L-57 to L-266; S-58 to L-266; C-59 toL-266; C-60 to L-266; L-61 to L-266; T-62 to L-266; V-63 to L-266; V-64to L-266; S-65 to L-266; F-66 to L-266; Y-67 to L-266; Q-68 to L-266;V-69 to L-266; A-70 to L-266; A-71 to L-266; L-72 to L-266; Q-73 toL-266; G-74 to L-266; D-75 to L-266; L-76 to L-266; A-77 to L-266; S-78to L-266; L-79 to L-266; R-80 to L-266; A-81 to L-266; E-82 to L-266;L-83 to L-266; Q-84 to L-266; G-85 to L-266; H-86 to L-266; H-87 toL-266; A-88 to L-266; E-89 to L-266; K-90 to L-266; L-91 to L-266; P-92to L-266; A-93 to L-266; G-94 to L-266; A-95 to L-266; G-96 to L-266;A-97 to L-266; P-98 to L-266; K-99 to L-266; A-100 to L-266; G-101 toL-266; L-102 to L-266; E-103 to L-266; E-104 to L-266; A-105 to L-266;P-106 to L-266; A-107 to L-266; V-108 to L-266; T-109 to L-266; A-110 toL-266; G-111 to L-266; L-112 to L-266; K-113 to L-266; I-114 to L-266;F-115 to L-266; E-116 to L-266; P-117 to L-266; P-118 to L-266; A-119 toL-266; P-120 to L-266; G-121 to L-266; E-122 to L-266; G-123 to L-266;N-124 to L-266; S-125 to L-266; S-126 to L-266; Q-127 to L-266; N-128 toL-266; S-129 to L-266; R-130 to L-266; N-131 to L-266; K-132 to L-266;R-133 to L-266; A-134 to L-266; V-135 to L-266; Q-136 to L-266; G-137 toL-266; P-138 to L-266; E-139 to L-266; E-140 to L-266; T-141 to L-266;G-142 to L-266; S-143 to L-266; Y-144 to L-266; T-145 to L-266; F-146 toL-266; V-147 to L-266; P-148 to L-266; W-149 to L-266; L-150 to L-266;L-151 to L-266; S-152 to L-266; F-153 to L-266; K-154 to L-266; R-155 toL-266; G-156 to L-266; S-157 to L-266; A-158 to L-266; L-159 to L-266;E-160 to L-266; E-161 to L-266; K-162 to L-266; E-163 to L-266; N-164 toL-266; K-165 to L-266; I-166 to L-266; L-167 to L-266; V-168 to L-266;K-169 to L-266; E-170 to L-266; T-171 to L-266; G-172 to L-266; Y-173 toL-266; F-174 to L-266; F-175 to L-266; I-176 to L-266; Y-177 to L-266;G-178 to L-266; Q-179 to L-266; V-180 to L-266; L-181 to L-266; Y-182 toL-266; T-183 to L-266; D-184 to L-266; K-185 to L-266; T-186 to L-266;Y-187 to L-266; A-188 to L-266; M-189 to L-266; G-190 to L-266; H-191 toL-266; L-192 to L-266; I-193 to L-266; Q-194 to L-266; R-195 to L-266;K-196 to L-266; K-197 to L-266; V-198 to L-266; H-199 to L-266; V-200 toL-266; F-201 to L-266; G-202 to L-266; D-203 to L-266; E-204 to L-266;L-205 to L-266; S-206 to L-266; L-207 to L-266; V-208 to L-266; T-209 toL-266; L-210 to L-266; F-211 to L-266; R-212 to L-266; C-213 to L-266;I-214 to L-266; Q-215 to L-266; N-216 to L-266; M-217 to L-266; P-218 toL-266; E-219 to L-266; T-220 to L-266; L-221 to L-266; P-222 to L-266;N-223 to L-266; N-224 to L-266; S-225 to L-266; C-226 to L-266; Y-227 toL-266; S-228 to L-266; A-229 to L-266; G-230 to L-266; I-231 to L-266;A-232 to L-266; K-233 to L-266; L-234 to L-266; E-235 to L-266; E-236 toL-266; G-237 to L-266; D-238 to L-266; E-239 to L-266; L-240 to L-266;Q-241 to L-266; L-242 to L-266; A-243 to L-266; I-244 to L-266; P-245 toL-266; R-246 to L-266; E-247 to L-266; N-248 to L-266; A-249 to L-266;Q-250 to L-266; I-251 to L-266; S-252 to L-266; L-253 to L-266; D-254 toL-266; G-255 to L-266; D-256 to L-266; V-257 to L-266; T-258 to L-266;F-259 to L-266; F-260 to L-266; and G-261 to L-266 of SEQ ID NO:3229.The present invention is also directed to antibodies that bind BLymphocyte Stimulator polypeptides comprising, or alternatively,consisting of, a contiguous sequence of amino acid residues at least80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99% identical to the aminoacid sequence of B Lymphocyte Stimulator polypeptides described above.

Also as mentioned above, even if deletion of one or more amino acidsfrom the C-terminus of a protein results in modification or loss of oneor more functional activities (e.g., biological activities) of theprotein, other functional activities may still be retained. Thus, theability of a shortened B Lymphocyte Stimulator mutein to induce and/orbind to antibodies which recognize the complete or mature form or theextracellular domain of the polypeptide generally will be retained whenless than the majority of the residues of the complete or mature form orthe extracellular domain of the polypeptide are removed from theC-terminus. Whether a particular polypeptide lacking C-terminal residuesof a complete polypeptide retains such immunologic activities canreadily be determined by routine methods described herein and otherwiseknown in the art. It is not unlikely that a B Lymphocyte Stimulatormutein with a large number of deleted C-terminal amino acid residues mayretain some functional (e.g., immunogenic) activities. In fact, peptidescomposed of as few as six B Lymphocyte Stimulator amino acid residuesmay often evoke an immune response.

Accordingly, the present invention further provides in anotherembodiment, antibodies that bind polypeptides having one or moreresidues deleted from the carboxy terminus of the amino acid sequence ofthe B Lymphocyte Stimulator shown in SEQ ID NO:3229, up to the glutamicacid residue at position number 6, and polynucleotides encoding suchpolypeptides. In particular, the present invention provides antibodiesthat bind polypeptides comprising the amino acid sequence of residues1-m⁵ of SEQ ID NO:3229, where m⁵ is an integer in the range of the aminoacid position of amino acid residues 6 to 265 in the amino acid sequenceof SEQ ID NO:3229.

More in particular, the invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues M-1 to L-265;M-1 to K-264; M-1 to L-263; M-1 to A-262; M-1 to G-261; M-1 to F-260;M-1 to F-259; M-1 to T-258; M-1 to V-257; M-1 to D-256; M-1 to G-255;M-1 to D-254; M-1 to L-253; M-1 to S-252; M-1 to I-251; M-1 to Q-250;M-1 to A-249; M-1 to N-248; M-1 to E-247; M-1 to R-246; M-1 to P-245;M-1 to I-244; M-1 to A-243; M-1 to L-242; M-1 to Q-241; M-1 to L-240;M-1 to E-239; M-1 to D-238; M-1 to G-237; M-1 to E-236; M-1 to E-235;M-1 to L-234; M-1 to K-233; M-1 to A-232; M-1 to I-231; M-1 to G-230;M-1 to A-229; M-1 to S-228; M-1 to Y-227; M-1 to C-226; M-1 to S-225;M-1 to N-224; M-1 to N-223; M-1 to P-222; M-1 to L-221; M-1 to T-220;M-1 to E-219; M-1 to P-218; M-1 to M-217; M-1 to N-216; M-1 to Q-215;M-1 to I-214; M-1 to C-213; M-1 to R-212; M-1 to F-211; M-1 to L-210;M-1 to T-209; M-1 to V-208; M-1 to L-207; M-1 to S-206; M-1 to L-205;M-1 to E-204; M-1 to D-203; M-1 to G-202; M-1 to F-201; M-1 to V-200;M-1 to H-199; M-1 to V-198; M-1 to K-197; M-1 to K-196; M-1 to R-195;M-1 to Q-194; M-1 to I-193; M-1 to L-192; M-1 to H-191; M-1 to G-190;M-1 to M-189; M-1 to A-188; M-1 to Y-187; M-1 to T-186; M-1 to K-185;M-1 to D-184; M-1 to T-183; M-1 to Y-182; M-1 to L-181; M-1 to V-180;M-1 to Q-179; M-1 to G-178; M-1 to Y-177; M-1 to I-176; M-1 to F-175;M-1 to F-174; M-1 to Y-173; M-1 to G-172; M-1 to T-171; M-1 to E-170;M-1 to K-169; M-1 to V-168; M-1 to L-167; M-1 to I-166; M-1 to K-165;M-1 to N-164; M-1 to E-163; M-1 to K-162; M-1 to E-161; M-1 to E-160;M-1 to L-159; M-1 to A-158; M-1 to S-157; M-1 to G-156; M-1 to R-155;M-1 to K-154; M-1 to F-153; M-1 to S-152; M-1 to L-151; M-1 to L-150;M-1 to W-149; M-1 to P-148; M-1 to V-147; M-1 to F-146; M-1 to T-145;M-1 to Y-144; M-1 to S-143; M-1 to G-142; M-1 to T-141; M-1 to E-140;M-1 to E-139; M-1 to P-138; M-1 to G-137; M-1 to Q-136; M-1 to V-135;M-1 to A-134; M-1 to R-133; M-1 to K-132; M-1 to N-131; M-1 to R-130;M-1 to S-129; M-1 to N-128; M-1 to Q-127; M-1 to S-126; M-1 to S-125;M-1 to N-124; M-1 to G-123; M-1 to E-122; M-1 to G-121; M-1 to P-120;M-1 to A-119; M-1 to P-118; M-1 to P-117; M-1 to E-116; M-1 to F-115;M-1 to I-114; M-1 to K-113; M-1 to L-112; M-1 to G-111; M-1 to A-110;M-1 to T-109; M-1 to V-108; M-1 to A-107; M-1 to P-106; M-1 to A-105;M-1 to E-104; M-1 to E-103; M-1 to L-102; M-1 to G-101; M-1 to A-100;M-1 to K-99; M-1 to P-98; M-1 to A-97; M-1 to G-96; M-1 to A-95; M-1 toG-94; M-1 to A-93; M-1 to P-92; M-1 to L-91; M-1 to K-90; M-1 to E-89;M-1 to A-88; M-1 to H-87; M-1 to H-86; M-1 to G-85; M-1 to Q-84; M-1 toL-83; M-1 to E-82; M-1 to A-81; M-1 to R-80; M-1 to L-79; M-1 to S-78;M-1 to A-77; M-1 to L-76; M-1 to D-75; M-1 to G-74; M-1 to Q-73; M-1 toL-72; M-1 to A-71; M-1 to A-70; M-1 to V-69; M-1 to Q-68; M-1 to Y-67;M-1 to F-66; M-1 to S-65; M-1 to V-64; M-1 to V-63; M-1 to T-62; M-1 toL-61; M-1 to C-60; M-1 to C-59; M-1 to S-58; M-1 to L-57; M-1 to L-56;M-1 to A-55; M-1 to L-54; M-1 to L-53; M-1 to L-52; M-1 to T-51; M-1 toA-50; M-1 to A-49; M-1 to L-48; M-1 to L-47; M-1 to K-46; M-1 to G-45;M-1 to D-44; M-1 to K-43; M-1 to S-42; M-1 to S-41; M-1 to R-40; M-1 toV-39; M-1 to S-38; M-1 to P-37; M-1 to S-36; M-1 to E-35; M-1 to K-34;M-1 to R-33; M-1 to P-32; M-1 to L-31; M-1 to I-30; M-1 to S-29; M-1 toV-28; M-1 to C-27; M-1 to E-26; M-1 to K-25; M-1 to L-24; M-1 to K-23;M-1 to M-22; M-1 to E-21; M-1 to E-20; M-1 to R-19; M-1 to K-18; M-1 toK-17; M-1 to L-16; M-1 to C-15; M-1 to S-14; M-1 to T-13; M-1 to L-12;M-1 to R-11; M-1 to S-10; M-1 to Q-9; M-1 to E-8; M-1 to R-7; and M-1 toE-6 of SEQ ID NO:3229. The present invention is also directed toantibodies that bind B Lymphocyte Stimulator polypeptides comprising, oralternatively, consisting of, a contiguous sequence of amino acidresidues at least 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99%identical to the amino acid sequence of B Lymphocyte Stimulatorpolypeptides described above.

The invention also provides antibodies that bind polypeptides having oneor more amino acids deleted from both the amino and the carboxyl terminiof a B Lymphocyte Stimulator polypeptide, which may be describedgenerally as having residues n⁵-m⁵ of SEQ ID NO:3229, where n⁵ and m⁵are integers as defined above.

In additional embodiments, the present invention provides antibodiesthat bind polypeptides comprising the amino acid sequence of residues134-m⁶ of SEQ ID NO:3228, where m⁶ is an integer from 140 to 285,corresponding to the position of the amino acid residue in SEQ IDNO:3228. For example, the invention provides antibodies that bindpolypeptides comprising, or alternatively consisting of, an amino acidsequence selected from the group consisting of residues A-134 toLeu-285; A-134 to L-284; A-134 to K-283; A-134 to L-282; A-134 to A-281;A-134 to G-280; A-134 to F-279; A-134 to F-278; A-134 to T-277; A-134 toV-276; A-134 to D-275; A-134 to G-274; A-134 to D-273; A-134 to L-272;A-134 to S-271; A-134 to I-270; A-134 to Q-269; A-134 to A-268; A-134 toN-267; A-134 to E-266; A-134 to R-265; A-134 to P-264; A-134 to I-263;A-134 to A-262; A-134 to L-261; A-134 to Q-260; A-134 to L-259; A-134 toE-258; A-134 to D-257; A-134 to G-256; A-134 to E-255; A-134 to E-254;A-134 to L-253; A-134 to K-252; A-134 to A-251; A-134 to I-250; A-134 toG-249; A-134 to A-248; A-134 to S-247; A-134 to Y-246; A-134 to C-245;A-134 to S-244; A-134 to N-243; A-134 to N-242; A-134 to P-241; A-134 toL-240; A-134 to T-239; A-134 to E-238; A-134 to P-237; A-134 to M-236;A-134 to N-235; A-134 to Q-234; A-134 to I-233; A-134 to C-232; A-134 toR-231; A-134 to F-230; A-134 to L-229; A-134 to T-228; A-134 to V-227;A-134 to L-226; A-134 to S-225; A-134 to L-224; A-134 to E-223; A-134 toD-222; A-134 to G-221; A-134 to F-220; A-134 to V-219; A-134 to H-218;A-134 to V-217; A-134 to K-216; A-134 to K-215; A-134 to R-214; A-134 toQ-213; A-134 to I-212; A-134 to L-211; A-134 to H-210; A-134 to G-209;A-134 to M-208; A-134 to A-207; A-134 to Y-206; A-134 to T-205; A-134 toK-204; A-134 to D-203; A-134 to T-202; A-134 to Y-201; A-134 to L-200;A-134 to V-199; A-134 to Q-198; A-134 to G-197; A-134 to Y-196; A-134 toI-195; A-134 to F-194; A-134 to F-193; A-134 to Y-192; A-134 to G-191;A-134 to T-190; A-134 to E-189; A-134 to K-188; A-134 to V-187; A-134 toL-186; A-134 to I-185; A-134 to K-184; A-134 to N-183; A-134 to E-182;A-134 to K-181; A-134 to E-180; A-134 to E-179; A-134 to L-178; A-134 toA-177; A-134 to S-176; A-134 to G-175; A-134 to R-174; A-134 to K-173;A-134 to F-172; A-134 to S-171; A-134 to L-170; A-134 to L-169; A-134 toW-168; A-134 to P-167; A-134 to V-166; A-134 to F-165; A-134 to T-164;A-134 to Y-163; A-134 to S-162; A-134 to G-161; A-134 to K-160; A-134 toQ-159; A-134 to I-158; A-134 to T-157; A-134 to P-156; A-134 to T-155;A-134 to E-154; A-134 to S-153; A-134 to D-152; A-134 to A-151; A-134 toI-150; A-134 to L-149; A-134 to Q-148; A-134 to L-147; A-134 to C-146;A-134 to D-145; A-134 to Q-144; A-134 to T-143; A-134 to V-142; A-134 toT-141; and A-134 to E-140 of SEQ ID NO:3228. The present invention isalso directed to antibodies that bind B Lymphocyte Stimulatorpolypeptides comprising, or alternatively, consisting of, a contiguoussequence of amino acid residues at least 80%, 85%, 90%, 92%, 95%, 96%,97%, 98% or 99% identical to the amino acid sequence of B LymphocyteStimulator polypeptides described above.

In additional embodiments, antibodies of the present invention may bindpolypeptide fragments comprising, or alternatively consisting of, anamino acid sequence selected from the group consisting of residues: M-1to C-15; D-2 to L-16; D-3 to K-17; S-4 to K-18; T-5 to R-19; E-6 toE-20; R-7 to E-21; E-8 to M-22; Q-9 to K-23; S-10 to L-24; R-11 to K-25;L-12 to E-26; T-13 to C-27; S-14 to V-28; C-15 to S-29; L-16 to I-30;K-17 to L-31; K-18 to P-32; R-19 to R-33; E-20 to K-34; E-21 to E-35;M-22 to S-36; K-23 to P-37; L-24 to S-38; K-25 to V-39; E-26 to R-40;C-27 to S-41; V-28 to S-42; S-29 to K-43; I-30 to D-44; L-31 to G-45;P-32 to K-46; R-33 to L-47; K-34 to L-48; E-35 to A-49; S-36 to A-50;P-37 to T-51; S-38 to L-52; V-39 to L-53; R-40 to L-54; S-41 to A-55;S-42 to L-56; K-43 to L-57; D-44 to S-58; G-45 to C-59; K-46 to C-60;L-47 to L-61; L-48 to T-62; A-49 to V-63; A-50 to V-64; T-51 to S-65;L-52 to F-66; L-53 to Y-67; L-54 to Q-68; A-55 to V-69; L-56 to A-70;L-57 to A-71; S-58 to L-72; C-59 to Q-73; C-60 to G-74; L-61 to D-75;T-62 to L-76; V-63 to A-77; V-64 to S-78; S-65 to L-79; F-66 to R-80;Y-67 to A-81; Q-68 to E-82; V-69 to L-83; A-70 to Q-84; A-71 to G-85;L-72 to H-86; Q-73 to H-87; G-74 to A-88; D-75 to E-89; L-76 to K-90;A-77 to L-91; S-78 to P-92; L-79 to A-93; R-80 to G-94; A-81 to A-95;E-82 to G-96; L-83 to A-97; Q-84 to P-98; G-85 to K-99; H-86 to A-100;H-87 to G-101; A-88 to L-102; E-89 to E-103; K-90 to E-104; L-91 toA-105; P-92 to P-106; A-93 to A-107; G-94 to V-108; A-95 to T-109; G-96to A-110; A-97 to G-111; P-98 to L-112; K-99 to K-113; A-100 to I-114;G-101 to F-115; L-102 to E-116; E-103 to P-117; E-104 to P-118; A-105 toA-119; P-106 to P-120; A-107 to G-121; V-108 to E-122; T-109 to G-123;A-110 to N-124; G-111 to S-125; L-112 to S-126; K-113 to Q-127; I-114 toN-128; F-115 to S-129; E-116 to R-130; P-117 to N-131; P-118 to K-132;A-119 to R-133; P-120 to A-134; G-121 to V-135; E-122 to Q-136; G-123 toG-137; N-124 to P-138; S-125 to E-139; S-126 to E-140; Q-127 to T-141;N-128 to V-142; S-129 to T-143; R-130 to Q-144; N-131 to D-145; K-132 toC-146; R-133 to L-147; A-134 to Q-148; V-135 to L-149; Q-136 to I-150;G-137 to A-151; P-138 to D-152; E-139 to S-153; E-140 to E-154; T-141 toT-155; V-142 to P-156; T-143 to T-157; Q-144 to I-158; D-145 to Q-159;C-146 to K-160; L-147 to G-161; Q-148 to S-162; L-149 to Y-163; I-150 toT-164; A-151 to F-165; D-152 to V-166; S-153 to P-167; E-154 to W-168;T-155 to L-169; P-156 to L-170; T-157 to S-171; I-158 to F-172; Q-159 toK-173; K-160 to R-174; G-161 to G-175; S-162 to S-176; Y-163 to A-177;T-164 to L-178; F-165 to E-179; V-166 to E-180; P-167 to K-181; W-168 toE-182; L-169 to N-183; L-170 to K-184; S-171 to I-185; F-172 to L-186;K-173 to V-187; R-174 to K-188; G-175 to E-189; S-176 to T-190; A-177 toG-191; L-178 to Y-192; E-179 to F-193; E-180 to F-194; K-181 to I-195;E-182 to Y-196; N-183 to G-197; K-184 to Q-198; I-185 to V-199; L-186 toL-200; V-187 to Y-201; K-188 to T-202; E-189 to D-203; T-190 to K-204;G-191 to T-205; Y-192 to Y-206; F-193 to A-207; F-194 to M-208; I-195 toG-209; Y-196 to H-210; G-197 to L-211; Q-198 to I-212; V-199 to Q-213;L-200 to R-214; Y-201 to K-215; T-202 to K-216; D-203 to V-217; K-204 toH-218; T-205 to V-219; Y-206 to F-220; A-207 to G-221; M-208 to D-222;G-209 to E-223; H-210 to L-224; L-211 to S-225; I-212 to L-226; Q-213 toV-227; R-214 to T-228; K-215 to L-229; K-216 to F-230; V-217 to R-231;H-218 to C-232; V-219 to I-233; F-220 to Q-234; G-221 to N-235; D-222 toM-236; E-223 to P-237; L-224 to E-238; S-225 to T-239; L-226 to L-240;V-227 to P-241; T-228 to N-242; L-229 to N-243; F-230 to S-244; R-231 toC-245; C-232 to Y-246; I-233 to S-247; Q-234 to A-248; N-235 to G-249;M-236 to I-250; P-237 to A-251; E-238 to K-252; T-239 to L-253; L-240 toE-254; P-241 to E-255; N-242 to G-256; N-243 to D-257; S-244 to E-258;C-245 to L-259; Y-246 to Q-260; S-247 to L-261; A-248 to A-262; G-249 toI-263; I-250 to P-264; A-251 to R-265; K-252 to E-266; L-253 to N-267;E-254 to A-268; E-255 to Q-269; G-256 to I-270; D-257 to S-271; E-258 toL-272; L-259 to D-273; Q-260 to G-274; L-261 to D-275; A-262 to V-276;I-263 to T-277; P-264 to F-278; R-265 to F-279; E-266 to G-280; N-267 toA-281; A-268 to L-282; Q-269 to K-283; I-270 to L-284; and S-271 toL-285 of SEQ ID NO:3228. The present invention is also directed toantibodies that bind B Lymphocyte Stimulator polypeptides comprising, oralternatively, consisting of, a contiguous sequence of amino acidresidues at least 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99%identical to the amino acid sequence of B Lymphocyte Stimulatorpolypeptides described above.

In additional embodiments, antibodies of the present invention may bindpolypeptide fragments comprising, or alternatively consisting of, anamino acid sequence selected from the group consisting of residues: M-1to C-15; D-2 to L-16; D-3 to K-17; S-4 to K-18; T-5 to R-19; E-6 toE-20; R-7 to E-21; E-8 to M-22; Q-9 to K-23; S-10 to L-24; R-11 to K-25;L-12 to E-26; T-13 to C-27; S-14 to V-28; C-15 to S-29; L-16 to I-30;K-17 to L-31; K-18 to P-32; R-19 to R-33; E-20 to K-34; E-21 to E-35;M-22 to S-36; K-23 to P-37; L-24 to S-38; K-25 to V-39; E-26 to R-40;C-27 to S-41; V-28 to S-42; S-29 to K-43; I-30 to D-44; L-31 to G-45;P-32 to K-46; R-33 to L-47; K-34 to L-48; E-35 to A-49; S-36 to A-50;P-37 to T-51; S-38 to L-52; V-39 to L-53; R-40 to L-54; S-41 to A-55;S-42 to L-56; K-43 to L-57; D-44 to S-58; G-45 to C-59; K-46 to C-60;L-47 to L-61; L-48 to T-62; A-49 to V-63; A-50 to V-64; T-51 to S-65;L-52 to F-66; L-53 to Y-67; L-54 to Q-68; A-55 to V-69; L-56 to A-70;L-57 to A-71; S-58 to L-72; C-59 to Q-73; C-60 to G-74; L-61 to D-75;T-62 to L-76; V-63 to A-77; V-64 to S-78; S-65 to L-79; F-66 to R-80;Y-67 to A-81; Q-68 to E-82; V-69 to L-83; A-70 to Q-84; A-71 to G-85;L-72 to H-86; Q-73 to H-87; G-74 to A-88; D-75 to E-89; L-76 to K-90;A-77 to L-91; S-78 to P-92; L-79 to A-93; R-80 to G-94; A-81 to A-95;E-82 to G-96; L-83 to A-97; Q-84 to P-98; G-85 to K-99; H-86 to A-100;H-87 to G-101; A-88 to L-102; E-89 to E-103; K-90 to E-104; L-91 toA-105; P-92 to P-106; A-93 to A-107; G-94 to V-108; A-95 to T-109; G-96to A-110; A-97 to G-111; P-98 to L-112; K-99 to K-113; A-100 to I-114;G-101 to F-115; L-102 to E-116; E-103 to P-117; E-104 to P-118; A-105 toA-119; P-106 to P-120; A-107 to G-121; V-108 to E-122; T-109 to G-123;A-110 to N-124; G-111 to S-125; L-112 to S-126; K-113 to Q-127; I-114 toN-128; F-115 to S-129; E-116 to R-130; P-117 to N-131; P-118 to K-132;A-119 to R-133; P-120 to A-134; G-121 to V-135; E-122 to Q-136; G-123 toG-137; N-124 to P-138; S-125 to E-139; S-126 to E-140; Q-127 to T-141;N-128 to G-142; S-129 to S-143; R-130 to Y-144; N-131 to T-145; K-132 toF-146; R-133 to V-147; A-134 to P-148; V-135 to W-149; Q-136 to L-150;G-137 to L-151; P-138 to S-152; E-139 to F-153; E-140 to K-154; T-141 toR-155; G-142 to G-156; S-143 to S-157; Y-144 to A-158; T-145 to L-159;F-146 to E-160; V-147 to E-161; P-148 to K-162; W-149 to E-163; L-150 toN-164; L-151 to K-165; S-152 to I-166; F-153 to L-167; K-154 to V-168;R-155 to K-169; G-156 to E-170; S-157 to T-171; A-158 to G-172; L-159 toY-173; E-160 to F-174; E-161 to F-175; K-162 to I-176; E-163 to Y-177;N-164 to G-178; K-165 to Q-179; I-166 to V-180; L-167 to L-181; V-168 toY-182; K-169 to T-183; E-170 to D-184; T-171 to K-185; G-172 to T-186;Y-173 to Y-187; F-174 to A-188; F-175 to M-189; I-176 to G-190; Y-177 toH-191; G-178 to L-192; Q-179 to I-193; V-180 to Q-194; L-181 to R-195;Y-182 to K-196; T-183 to K-197; D-184 to V-198; K-185 to H-199; T-186 toV-200; Y-187 to F-201; A-188 to G-202; M-189 to D-203; G-190 to E-204;H-191 to L-205; L-192 to S-206; I-193 to L-207; Q-194 to V-208; R-195 toT-209; K-196 to L-210; K-197 to F-211; V-198 to R-212; H-199 to C-213;V-200 to I-214; F-201 to Q-215; G-202 to N-216; D-203 to M-217; E-204 toP-218; L-205 to E-219; S-206 to T-220; L-207 to L-221; V-208 to P-222;T-209 to N-223; L-210 to N-224; F-211 to S-225; R-212 to C-226; C-213 toY-227; I-214 to S-228; Q-215 to A-229; N-216 to G-230; M-217 to I-231;P-218 to A-232; E-219 to K-233; T-220 to L-234; L-221 to E-235; P-222 toE-236; N-223 to G-237; N-224 to D-238; S-225 to E-239; C-226 to L-240;Y-227 to Q-241; S-228 to L-242; A-229 to A-243; G-230 to I-244; I-231 toP-245; A-232 to R-246; K-233 to E-247; L-234 to N-248; E-235 to A-249;E-236 to Q-250; G-237 to I-251; D-238 to S-252; E-239 to L-253; L-240 toD-254; Q-241 to G-255; L-242 to D-256; A-243 to V-257; I-244 to T-258;P-245 to F-259; R-246 to F-260; E-247 to G-261; N-248 to A-262; A-249 toL-263; Q-250 to K-264; I-251 to L-265; and S-252 to L-266 of SEQ IDNO:3229. The present invention is also directed to antibodies that bindB Lymphocyte Stimulator polypeptides comprising, or alternatively,consisting of, a contiguous sequence of amino acid residues at least80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99% identical to the aminoacid sequence of B Lymphocyte Stimulator polypeptides described above.

In additional embodiments, antibodies of the present invention may bindpolypeptide fragments comprising, or alternatively consisting of, anamino acid sequence selected from the group consisting of residues: M-1to F-15; D-2 to C-16; E-3 to S-17; S-4 to E-18; A-5 to K-19; K-6 toG-20; T-7 to E-21; L-8 to D-22; P-9 to M-23; P-10 to K-24; P-11 to V-25;C-12 to G-26; L-13 to Y-27; C-14 to D-28; F-15 to P-29; C-16 to I-30;S-17 to T-31; E-18 to P-32; K-19 to Q-33; G-20 to K-34; E-21 to E-35;D-22 to E-36; M-23 to G-37; K-24 to A-38; V-25 to W-39; G-26 to F-40;Y-27 to G-41; D-28 to I-42; P-29 to C-43; I-30 to R-44; T-31 to D-45;P-32 to G-46; Q-33 to R-47; K-34 to L-48; E-35 to L-49; E-36 to A-50;G-37 to A-51; A-38 to T-52; W-39 to L-53; F-40 to L-54; G-41 to L-55;I-42 to A-56; C-43 to L-57; R-44 to L-58; D-45 to S-59; G-46 to S-60;R-47 to S-61; L-48 to F-62; L-49 to T-63; A-50 to A-64; A-51 to M-65;T-52 to S-66; L-53 to L-67; L-54 to Y-68; L-55 to Q-69; A-56 to L-70;L-57 to A-71; L-58 to A-72; S-59 to L-73; S-60 to Q-74; S-61 to A-75;F-62 to D-76; T-63 to L-77; A-64 to M-78; M-65 to N-79; S-66 to L-80;L-67 to R-81; Y-68 to M-82; Q-69 to E-83; L-70 to L-84; A-71 to Q-85;A-72 to S-86; L-73 to Y-87; Q-74 to R-88; A-75 to G-89; D-76 to S-90;L-77 to A-91; M-78 to T-92; N-79 to P-93; L-80 to A-94; R-81 to A-95;M-82 to A-96; E-83 to G-97; L-84 to A-98; Q-85 to P-99; S-86 to E-100;Y-87 to L-101; R-88 to T-102; G-89 to A-103; S-90 to G-104; A-91 toV-105; T-92 to K-106; P-93 to L-107; A-94 to L-108; A-95 to T-109; A-96to P-110; G-97 to A-111; A-98 to A-112; P-99 to P-113; E-100 to R-114;L-101 to P-115; T-102 to H-116; A-103 to N-117; G-104 to S-118; V-105 toS-119; K-106 to R-120; L-107 to G-121; L-108 to H-122; T-109 to R-123;P-110 to N-124; A-111 to R-125; A-112 to R-126; P-113 to A-127; R-114 toF-128; P-115 to Q-129; H-116 to G-130; N-117 to P-131; S-118 to E-132;S-119 to E-133; R-120 to T-134; G-121 to E-135; H-122 to Q-136; R-123 toD-137; N-124 to V-138; R-125 to D-139; R-126 to L-140; A-127 to S-141;F-128 to A-142; Q-129 to P-143; G-130 to P-144; P-131 to A-145; E-132 toP-146; E-133 to C-147; T-134 to L-148; E-135 to P-149; Q-136 to G-150;D-137 to C-151; V-138 to R-152; D-139 to H-153; L-140 to S-154; S-141 toQ-155; A-142 to H-156; P-143 to D-157; P-144 to D-158; A-145 to N-159;P-146 to G-160; C-147 to M-161; L-148 to N-162; P-149 to L-163; G-150 toR-164; C-151 to N-165; R-152 to I-166; H-153 to I-167; S-154 to Q-168;Q-155 to D-169; H-156 to C-170; D-157 to L-171; D-158 to Q-172; N-159 toL-173; G-160 to I-174; M-161 to A-175; N-162 to D-176; L-163 to S-177;R-164 to D-178; N-165 to T-179; I-166 to P-180; I-167 to A-181; Q-168 toL-182; D-169 to E-183; C-170 to E-184; L-171 to K-185; Q-172 to E-186;L-173 to N-187; I-174 to K-188; A-175 to I-189; D-176 to V-190; S-177 toV-191; D-178 to R-192; T-179 to Q-193; P-180 to T-194; A-181 to G-195;L-182 to Y-196; E-183 to F-197; E-184 to F-198; K-185 to I-199; E-186 toY-200; N-187 to S-201; K-188 to Q-202; I-189 to V-203; V-190 to L-204;V-191 to Y-205; R-192 to T-206; Q-193 to D-207; T-194 to P-208; G-195 toI-209; Y-196 to F-210; F-197 to A-211; F-198 to M-212; I-199 to G-213;Y-200 to H-214; S-201 to V-215; Q-202 to I-216; V-203 to Q-217; L-204 toR-218; Y-205 to K-219; T-206 to K-220; D-207 to V-221; P-208 to H-222;I-209 to V-223; F-210 to F-224; A-211 to G-225; M-212 to D-226; G-213 toE-227; H-214 to L-228; V-215 to S-229; I-216 to L-230; Q-217 to V-231;R-218 to T-232; K-219 to L-233; K-220 to F-234; V-221 to R-235; H-222 toC-236; V-223 to I-237; F-224 to Q-238; G-225 to N-239; D-226 to M-240;E-227 to P-241; L-228 to K-242; S-229 to T-243; L-230 to L-244; V-231 toP-245; T-232 to N-246; L-233 to N-247; F-234 to S-248; R-235 to C-249;C-236 to Y-250; I-237 to S-251; Q-238 to A-252; N-239 to G-253; M-240 toI-254; P-241 to A-255; K-242 to R-256; T-243 to L-257; L-244 to E-258;P-245 to E-259; N-246 to G-260; N-247 to D-261; S-248 to E-262; C-249 toI-263; Y-250 to Q-264; S-251 to L-265; A-252 to A-266; G-253 to I-267;I-254 to P-268; A-255 to R-269; R-256 to E-270; L-257 to N-271; E-258 toA-272; E-259 to Q-273; G-260 to I-274; D-261 to S-275; E-262 to R-276;I-263 to N-277; Q-264 to G-278; L-265 to D-279; A-266 to D-280; I-267 toT-281; P-268 to F-282; R-269 to F-283; E-270 to G-284; N-271 to A-285;A-272 to L-286; Q-273 to K-287; I-274 to L-288; and S-275 to L-289 ofSEQ ID NO:38. The present invention is also directed to antibodies thatbind B Lymphocyte Stimulator polypeptides comprising, or alternatively,consisting of, a contiguous sequence of amino acid residues at least80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99% identical to the aminoacid sequence of B Lymphocyte Stimulator polypeptides described above.

It will be recognized by one of ordinary skill in the art that someamino acid sequences of the B Lymphocyte Stimulator polypeptides can bevaried without significant effect of the structure or function of thepolypeptide. If such differences in sequence are contemplated, it shouldbe remembered that there will be critical areas on the polypeptide whichdetermine activity.

Thus, the invention further includes antibodies that bind variations ofB Lymphocyte Stimulator polypeptides which show B Lymphocyte Stimulatorpolypeptide functional activity (e.g., biological activity) or whichinclude regions of B Lymphocyte Stimulator polypeptide such as thepolypeptide fragments described herein. Such mutants include deletions,insertions, inversions, repeats, and type substitutions selectedaccording to general rules known in the art so as have little effect onactivity. For example, guidance concerning how to make phenotypicallysilent amino acid substitutions is provided in Bowie, J. U. et al.,“Deciphering the Message in Protein Sequences: Tolerance to Amino AcidSubstitutions,” Science 247:1306-1310 (1990), wherein the authorsindicate that there are two main approaches for studying the toleranceof an amino acid sequence to change. The first method relies on theprocess of evolution, in which mutations are either accepted or rejectedby natural selection. The second approach uses genetic engineering tointroduce amino acid changes at specific positions of a cloned gene andselections or screens to identify sequences that maintain functionality.

As the authors state, these studies have revealed that proteins aresurprisingly tolerant of amino acid substitutions. The authors furtherindicate which amino acid changes are likely to be permissive at acertain position of the protein. For example, most buried amino acidresidues require nonpolar side chains, whereas few features of surfaceside chains are generally conserved. Other such phenotypically silentsubstitutions are described in Bowie, J. U. et al., supra, and thereferences cited therein. Typically seen as conservative substitutionsare the replacements, one for another, among the aliphatic amino acidsAla, Val, Leu and Ile; interchange of the hydroxyl residues Ser and Thr,exchange of the acidic residues Asp and Glu, substitution between theamide residues Asn and Gln, exchange of the basic residues Lys and Argand replacements among the aromatic residues Phe, Tyr.

Thus, antibodies of the present invention may bind fragments,derivatives or analogs of the polypeptide of SEQ ID NO:3228, or thatencoded by the deposited cDNA plasmid, such as (i) polypeptides in whichone or more of the amino acid residues are substituted with a conservedor non-conserved amino acid residue (preferably a conserved amino acidresidue) and such substituted amino acid residue may or may not be oneencoded by the genetic code, or (ii) polypeptides in which one or moreof the amino acid residues includes a substituent group, or (iii)polypeptides in which the extracellular domain of the polypeptide isfused with another compound, such as a compound to increase thehalf-life of the polypeptide (for example, polyethylene glycol), or (iv)polypeptides in which the additional amino acids are fused to theextracellular domain of the polypeptide, such as an IgG Fc fusion regionpeptide or leader or secretory sequence or a sequence which is employedfor purification of the extracellular domain of the polypeptide or aproprotein sequence.

Antibodies of the present invention may bind fragments, derivatives oranalogs of the polypeptide of SEQ ID NO:3229, or that encoded by thedeposited cDNA plasmid, such as (i) polypeptides in which one or more ofthe amino acid residues are substituted with a conserved ornon-conserved amino acid residue (preferably a conserved amino acidresidue) and such substituted amino acid residue may or may not be oneencoded by the genetic code, or (ii) polypeptides in which one or moreof the amino acid residues includes a substituent group, or (iii)polypeptides in which the extracellular domain of the polypeptide isfused with another compound, such as a compound to increase thehalf-life of the polypeptide (for example, polyethylene glycol), or (iv)polypeptides in which the additional amino acids are fused to theextracellular domain of the polypeptide, such as, a soluble biologicallyactive fragment of another TNF ligand family member (e.g., CD40 Ligand),an IgG Fc fusion region peptide or leader or secretory sequence or asequence which is employed for purification of the extracellular domainof the polypeptide or a proprotein sequence. Such fragments, derivativesand analogs are deemed to be within the scope of those skilled in theart from the teachings herein.

Thus, the antibodies of the invention may bind B Lymphocyte Stimulatorpolypeptides that include one or more amino acid substitutions,deletions or additions, either from natural mutations or humanmanipulation. As indicated, changes are preferably of a minor nature,such as conservative amino acid substitutions that do not significantlyaffect the folding or activity of the protein (see Table 13).

TABLE 13 Conservative Amino Acid Substitutions. Aromatic PhenylalanineTryptophan Tyrosine Hydrophobic Leucine Isoleucine Valine PolarGlutamine Asparagine Basic Arginine Lysine Histidine Acidic AsparticAcid Glutamic Acid Small Alanine Serine Threonine Methionine Glycine

In one embodiment of the invention, antibodies of the present inventionbind polypeptides comprising, or alternatively consisting of, the aminoacid sequence of a B Lymphocyte Stimulator polypeptide having an aminoacid sequence which contains at least one conservative amino acidsubstitution, but not more than 50 conservative amino acidsubstitutions, even more preferably, not more than 40 conservative aminoacid substitutions, still more preferably, not more than 30 conservativeamino acid substitutions, and still even more preferably, not more than20 conservative amino acid substitutions. In one embodiment of theinvention, antibodies of the present invention bind polypeptidescomprising, or alternatively consisting of, the amino acid sequence of aB Lymphocyte Stimulator polypeptide having an amino acid sequence whichcontains at least one conservative amino acid substitution, but not morethan 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 conservative amino acidsubstitutions.

For example, site directed changes at the amino acid level of BLymphocyte Stimulator can be made by replacing a particular amino acidwith a conservative substitution. Antibodies of the present inventionmay bind B Lymphocyte Stimulator amino acid sequences containingconservative substitution mutations of the polypeptide of SEQ ID NO:3228including: M1 replaced with A, G, I, L, S, T, or V; D2 replaced with E;D3 replaced with E; S4 replaced with A, G, I, L, T, M, or V; T5 replacedwith A, G, I, L, S, M, or V; E6 replaced with D; R7 replaced with H, orK; E8 replaced with D; Q9 replaced with N; S10 replaced with A, G, I, L,T, M, or V; R11 replaced with H, or K; L12 replaced with A, G, I, S, T,M, or V; T13 replaced with A, G, I, L, S, M, or V; S14 replaced with A,G, I, L, T, M, or V; L16 replaced with A, G, I, S, T, M, or V; K17replaced with H, or R; K18 replaced with H, or R; R19 replaced with H,or K; E20 replaced with D; E21 replaced with D; M22 replaced with A, G,I, L, S, T, or V; K23 replaced with H, or R; L24 replaced with A, G, I,S, T, M, or V; K25 replaced with H, or R; E26 replaced with D; V28replaced with A, G, I, L, S, T, or M; S29 replaced with A, G, I, L, T,M, or V; I30 replaced with A, G, L, S, T, M, or V; L31 replaced with A,G, I, S, T, M, or V; R33 replaced with H, or K; K34 replaced with H, orR; E35 replaced with D; S36 replaced with A, G, I, L, T, M, or V; S38replaced with A, G, I, L, T, M, or V; V39 replaced with A, G, I, L, S,T, or M; R40 replaced with H, or K; S41 replaced with A, G, I, L, T, M,or V; S42 replaced with A, G, I, L, T, M, or V; K43 replaced with H, orR; D44 replaced with E; G45 replaced with A, I, L, S, T, M, or V; K46replaced with H, or R; L47 replaced with A, G, I, S, T, M, or V; L48replaced with A, G, I, S, T, M, or V; A49 replaced with G, I, L, S, T,M, or V; A50 replaced with G, I, L, S, T, M, or V; T51 replaced with A,G, I, L, S, M, or V; L52 replaced with A, G, I, S, T, M, or V; L53replaced with A, G, I, S, T, M, or V; L54 replaced with A, G, I, S, T,M, or V; A55 replaced with G, I, L, S, T, M, or V; L56 replaced with A,G, I, S, T, M, or V; L57 replaced with A, G, I, S, T, M, or V; S58replaced with A, G, I, L, T, M, or V; L61 replaced with A, G, I, S, T,M, or V; T62 replaced with A, G, I, L, S, M, or V; V63 replaced with A,G, I, L, S, T, or M; V64 replaced with A, G, I, L, S, T, or M; S65replaced with A, G, I, L, T, M, or V; F66 replaced with W, or Y; Y67replaced with F, or W; Q68 replaced with N; V69 replaced with A, G, I,L, S, T, or M; A70 replaced with G, I, L, S, T, M, or V; A71 replacedwith G, I, L, S, T, M, or V; L72 replaced with A, G, I, S, T, M, or V;Q73 replaced with N; G74 replaced with A, I, L, S, T, M, or V; D75replaced with E; L76 replaced with A, G, I, S, T, M, or V; A77 replacedwith G, I, L, S, T, M, or V; S78 replaced with A, G, I, L, T, M, or V;L79 replaced with A, G, I, S, T, M, or V; R80 replaced with H, or K; A81replaced with G, I, L, S, T, M, or V; E82 replaced with D; L83 replacedwith A, G, I, S, T, M, or V; Q84 replaced with N; G85 replaced with A,I, L, S, T, M, or V; H86 replaced with K, or R; H87 replaced with K, orR; A88 replaced with G, I, L, S, T, M, or V; E89 replaced with D; K90replaced with H, or R; L91 replaced with A, G, I, S, T, M, or V; A93replaced with G, I, L, S, T, M, or V; G94 replaced with A, I, L, S, T,M, or V; A95 replaced with G, I, L, S, T, M, or V; G96 replaced with A,I, L, S, T, M, or V; A97 replaced with G, I, L, S, T, M, or V; K99replaced with H, or R; A100 replaced with G, I, L, S, T, M, or V; G101replaced with A, I, L, S, T, M, or V; L102 replaced with A, G, I, S, T,M, or V; E103 replaced with D; E104 replaced with D; A105 replaced withG, I, L, S, T, M, or V; A107 replaced with G, I, L, S, T, M, or V; V108replaced with A, G, I, L, S, T, or M; T109 replaced with A, G, I, L, S,M, or V; A110 replaced with G, I, L, S, T, M, or V; G111 replaced withA, I, L, S, T, M, or V; L112 replaced with A, G, I, S, T, M, or V; K113replaced with H, or R; 1114 replaced with A, G, L, S, T, M, or V; F115replaced with W, or Y; E116 replaced with D; A119 replaced with G, I, L,S, T, M, or V; G121 replaced with A, I, L, S, T, M, or V; E122 replacedwith D; G123 replaced with A, I, L, S, T, M, or V; N124 replaced with Q;S125 replaced with A, G, I, L, T, M, or V; S126 replaced with A, G, I,L, T, M, or V; Q127 replaced with N; N128 replaced with Q; S129 replacedwith A, G, I, L, T, M, or V; R130 replaced with H, or K; N131 replacedwith Q; K132 replaced with H, or R; R133 replaced with H, or K; A134replaced with G, I, L, S, T, M, or V; V135 replaced with A, G, I, L, S,T, or M; Q136 replaced with N; G137 replaced with A, I, L, S, T, M, orV; E139 replaced with D; E140 replaced with D; T141 replaced with A, G,I, L, S, M, or V; V142 replaced with A, G, I, L, S, T, or M; T143replaced with A, G, I, L, S, M, or V; Q144 replaced with N; D145replaced with E; L147 replaced with A, G, I, S, T, M, or V; Q148replaced with N; L149 replaced with A, G, I, S, T, M, or V; I150replaced with A, G, L, S, T, M, or V; A151 replaced with G, I, L, S, T,M, or V; D152 replaced with E; S153 replaced with A, G, I, L, T, M, orV; E154 replaced with D; T155 replaced with A, G, I, L, S, M, or V; T157replaced with A, G, I, L, S, M, or V; I158 replaced with A, G, L, S, T,M, or V; Q159 replaced with N; K160 replaced with H, or R; G161 replacedwith A, I, L, S, T, M, or V; S162 replaced with A, G, I, L, T, M, or V;Y163 replaced with F, or W; T164 replaced with A, G, I, L, S, M, or V;F165 replaced with W, or Y; V166 replaced with A, G, I, L, S, T, or M;W168 replaced with F, or Y; L169 replaced with A, G, I, S, T, M, or V;L170 replaced with A, G, I, S, T, M, or V; S171 replaced with A, G, I,L, T, M, or V; F172 replaced with W, or Y; K173 replaced with H, or R;R174 replaced with H, or K; G175 replaced with A, I, L, S, T, M, or V;S176 replaced with A, G, I, L, T, M, or V; A177 replaced with G, I, L,S, T, M, or V; L178 replaced with A, G, I, S, T, M, or V; E179 replacedwith D; E180 replaced with D; K181 replaced with H, or R; E182 replacedwith D; N183 replaced with Q; K184 replaced with H, or R; I185 replacedwith A, G, L, S, T, M, or V; L186 replaced with A, G, I, S, T, M, or V;V187 replaced with A, G, I, L, S, T, or M; K188 replaced with H, or R;E189 replaced with D; T190 replaced with A, G, I, L, S, M, or V; G191replaced with A, I, L, S, T, M, or V; Y192 replaced with F, or W; F193replaced with W, or Y; F194 replaced with W, or Y; I195 replaced with A,G, L, S, T, M, or V; Y196 replaced with F, or W; G197 replaced with A,I, L, S, T, M, or V; Q198 replaced with N; V199 replaced with A, G, I,L, S, T, or M; L200 replaced with A, G, I, S, T, M, or V; Y201 replacedwith F, or W; T202 replaced with A, G, I, L, S, M, or V; D203 replacedwith E; K204 replaced with H, or R; T205 replaced with A, G, I, L, S, M,or V; Y206 replaced with F, or W; A207 replaced with G, I, L, S, T, M,or V; M208 replaced with A, G, I, L, S, T, or V; G209 replaced with A,I, L, S, T, M, or V; H210 replaced with K, or R; L211 replaced with A,G, I, S, T, M, or V; I212 replaced with A, G, L, S, T, M, or V; Q213replaced with N; 8214 replaced with H, or K; K215 replaced with H, or R;K216 replaced with H, or R; V217 replaced with A, G, I, L, S, T, or M;H218 replaced with K, or R; V219 replaced with A, G, I, L, S, T, or M;F220 replaced with W, or Y; G221 replaced with A, I, L, S, T, M, or V;D222 replaced with E; E223 replaced with D; L224 replaced with A, G, I,S, T, M, or V; S225 replaced with A, G, I, L, T, M, or V; L226 replacedwith A, G, I, S, T, M, or V; V227 replaced with A, G, I, L, S, T, or M;T228 replaced with A, G, I, L, S, M, or V; L229 replaced with A, G, I,S, T, M, or V; F230 replaced with W, or Y; R231 replaced with H, or K;I233 replaced with A, G, L, S, T, M, or V; Q234 replaced with N; N235replaced with Q; M236 replaced with A, G, I, L, S, T, or V; E238replaced with D; T239 replaced with A, G, I, L, S, M, or V; L240replaced with A, G, I, S, T, M, or V; N242 replaced with Q; N243replaced with Q; S244 replaced with A, G, I, L, T, M, or V; Y246replaced with F, or W; S247 replaced with A, G, I, L, T, M, or V; A248replaced with G, I, L, S, T, M, or V; G249 replaced with A, I, L, S, T,M, or V; I250 replaced with A, G, L, S, T, M, or V; A251 replaced withG, I, L, S, T, M, or V; K252 replaced with H, or R; L253 replaced withA, G, I, S, T, M, or V; E254 replaced with D; E255 replaced with D; G256replaced with A, I, L, S, T, M, or V; D257 replaced with E; E258replaced with D; L259 replaced with A, G, I, S, T, M, or V; Q260replaced with N; L261 replaced with A, G, I, S, T, M, or V; A262replaced with G, I, L, S, T, M, or V; I263 replaced with A, G, L, S, T,M, or V; R265 replaced with H, or K; E266 replaced with D; N267 replacedwith Q; A268 replaced with G, I, L, S, T, M, or V; Q269 replaced with N;I270 replaced with A, G, L, S, T, M, or V; S271 replaced with A, G, I,L, T, M, or V; L272 replaced with A, G, I, S, T, M, or V; D273 replacedwith E; G274 replaced with A, I, L, S, T, M, or V; D275 replaced with E;V276 replaced with A, G, I, L, S, T, or M; T277 replaced with A, G, I,L, S, M, or V; F278 replaced with W, or Y; F279 replaced with W, or Y;G280 replaced with A, I, L, S, T, M, or V; A281 replaced with G, I, L,S, T, M, or V; L282 replaced with A, G, I, S, T, M, or V; K283 replacedwith H, or R; L284 replaced with A, G, I, S, T, M, or V; and/or L285replaced with A, G, I, S, T, M, or V.

In another embodiment, site directed changes at the amino acid level ofB Lymphocyte Stimulator can be made by replacing a particular amino acidwith a conservative substitution. Antibodies of the present inventionmay bind B Lymphocyte Stimulator amino acid sequences containingconservative substitution mutations of the polypeptide of SEQ ID NO:3229including: M1 replaced with A, G, I, L, S, T, or V; D2 replaced with E;D3 replaced with E; S4 replaced with A, G, I, L, T, M, or V; T5 replacedwith A, G, I, L, S, M, or V; E6 replaced with D; R7 replaced with H, orK; E8 replaced with D; Q9 replaced with N; S10 replaced with A, G, I, L,T, M, or V; R11 replaced with H, or K; L12 replaced with A, G, I, S, T,M, or V; T13 replaced with A, G, I, L, S, M, or V; S14 replaced with A,G, I, L, T, M, or V; L16 replaced with A, G, I, S, T, M, or V; K17replaced with H, or R; K18 replaced with H, or R; R19 replaced with H,or K; E20 replaced with D; E21 replaced with D; M22 replaced with A, G,I, L, S, T, or V; K23 replaced with H, or R; L24 replaced with A, G, I,S, T, M, or V; K25 replaced with H, or R; E26 replaced with D; V28replaced with A, G, I, L, S, T, or M; S29 replaced with A, G, I, L, T,M, or V; I30 replaced with A, G, L, S, T, M, or V; L31 replaced with A,G, I, S, T, M, or V; R33 replaced with H, or K; K34 replaced with H, orR; E35 replaced with D; S36 replaced with A, G, I, L, T, M, or V; S38replaced with A, G, I, L, T, M, or V; V39 replaced with A, G, I, L, S,T, or M; R40 replaced with H, or K; S41 replaced with A, G, I, L, T, M,or V; S42 replaced with A, G, I, L, T, M, or V; K43 replaced with H, orR; D44 replaced with E; G45 replaced with A, I, L, S, T, M, or V; K46replaced with H, or R; L47 replaced with A, G, I, S, T, M, or V; L48replaced with A, G, I, S, T, M, or V; A49 replaced with G, I, L, S, T,M, or V; A50 replaced with G, I, L, S, T, M, or V; T51 replaced with A,G, I, L, S, M, or V; L52 replaced with A, G, I, S, T, M, or V; L53replaced with A, G, I, S, T, M, or V; L54 replaced with A, G, I, S, T,M, or V; A55 replaced with G, I, L, S, T, M, or V; L56 replaced with A,G, I, S, T, M, or V; L57 replaced with A, G, I, S, T, M, or V; S58replaced with A, G, I, L, T, M, or V; L61 replaced with A, G, I, S, T,M, or V; T62 replaced with A, G, I, L, S, M, or V; V63 replaced with A,G, I, L, S, T, or M; V64 replaced with A, G, I, L, S, T, or M; S65replaced with A, G, I, L, T, M, or V; F66 replaced with W, or Y; Y67replaced with F, or W; Q68 replaced with N; V69 replaced with A, G, I,L, S, T, or M; A70 replaced with G, I, L, S, T, M, or V; A71 replacedwith G, I, L, S, T, M, or V; L72 replaced with A, G, I, S, T, M, or V;Q73 replaced with N; G74 replaced with A, I, L, S, T, M, or V; D75replaced with E; L76 replaced with A, G, I, S, T, M, or V; A77 replacedwith G, I, L, S, T, M, or V; S78 replaced with A, G, I, L, T, M, or V;L79 replaced with A, G, I, S, T, M, or V; R80 replaced with H, or K; A81replaced with G, I, L, S, T, M, or V; E82 replaced with D; L83 replacedwith A, G, I, S, T, M, or V; Q84 replaced with N; G85 replaced with A,I, L, S, T, M, or V; H86 replaced with K, or R; H87 replaced with K, orR; A88 replaced with G, I, L, S, T, M, or V; E89 replaced with D; K90replaced with H, or R; L91 replaced with A, G, I, S, T, M, or V; A93replaced with G, I, L, S, T, M, or V; G94 replaced with A, I, L, S, T,M, or V; A95 replaced with G, I, L, S, T, M, or V; G96 replaced with A,I, L, S, T, M, or V; A97 replaced with G, I, L, S, T, M, or V; K99replaced with H, or R; A100 replaced with G, I, L, S, T, M, or V; G101replaced with A, I, L, S, T, M, or V; L102 replaced with A, G, I, S, T,M, or V; E103 replaced with D; E104 replaced with D; A105 replaced withG, I, L, S, T, M, or V; A107 replaced with G, I, L, S, T, M, or V; V108replaced with A, G, I, L, S, T, or M; T109 replaced with A, G, I, L, S,M, or V; A110 replaced with G, I, L, S, T, M, or V; G111 replaced withA, I, L, S, T, M, or V; L112 replaced with A, G, I, S, T, M, or V; K113replaced with H, or R; I114 replaced with A, G, L, S, T, M, or V; F115replaced with W, or Y; E116 replaced with D; A119 replaced with G, I, L,S, T, M, or V; G121 replaced with A, I, L, S, T, M, or V; E122 replacedwith D; G123 replaced with A, I, L, S, T, M, or V; N124 replaced with Q;S125 replaced with A, G, I, L, T, M, or V; S126 replaced with A, G, I,L, T, M, or V; Q127 replaced with N; N128 replaced with Q; S129 replacedwith A, G, I, L, T, M, or V; R130 replaced with H, or K; N131 replacedwith Q; K132 replaced with H, or R; R133 replaced with H, or K; A134replaced with G, I, L, S, T, M, or V; V135 replaced with A, G, I, L, S,T, or M; Q136 replaced with N; G137 replaced with A, I, L, S, T, M, orV; E139 replaced with D; E140 replaced with D; T141 replaced with A, G,I, L, S, M, or V; G142 replaced with A, I, L, S, T, M, or V; S143replaced with A, G, I, L, T, M, or V; Y144 replaced with F, or W; T145replaced with A, G, I, L, S, M, or V; F146 replaced with W, or Y; V147replaced with A, G, I, L, S, T, or M; W149 replaced with F, or Y; L150replaced with A, G, I, S, T, M, or V; L151 replaced with A, G, I, S, T,M, or V; S152 replaced with A, G, I, L, T, M, or V; F153 replaced withW, or Y; K154 replaced with H, or R; R155 replaced with H, or K; G156replaced with A, I, L, S, T, M, or V; S157 replaced with A, G, I, L, T,M, or V; A158 replaced with G, I, L, S, T, M, or V; L159 replaced withA, G, I, S, T, M, or V; E160 replaced with D; E161 replaced with D; K162replaced with H, or R; E163 replaced with D; N164 replaced with Q; K165replaced with H, or R; I166 replaced with A, G, L, S, T, M, or V; L167replaced with A, G, I, S, T, M, or V; V168 replaced with A, G, I, L, S,T, or M; K169 replaced with H, or R; E170 replaced with D; T171 replacedwith A, G, I, L, S, M, or V; G172 replaced with A, I, L, S, T, M, or V;Y173 replaced with F, or W; F174 replaced with W, or Y; F175 replacedwith W, or Y; I176 replaced with A, G, L, S, T, M, or V; Y177 replacedwith F, or W; G178 replaced with A, I, L, S, T, M, or V; Q179 replacedwith N; V180 replaced with A, G, I, L, S, T, or M; L181 replaced with A,G, I, S, T, M, or V; Y182 replaced with F, or W; T183 replaced with A,G, I, L, S, M, or V; D184 replaced with E; K185 replaced with H, or R;T186 replaced with A, G, I, L, S, M, or V; Y187 replaced with F, or W;A188 replaced with G, I, L, S, T, M, or V; M189 replaced with A, G, I,L, S, T, or V; G190 replaced with A, I, L, S, T, M, or V; H191 replacedwith K, or R; L192 replaced with A, G, I, S, T, M, or V; I193 replacedwith A, G, L, S, T, M, or V; Q194 replaced with N; R195 replaced with H,or K; K196 replaced with H, or R; K197 replaced with H, or R; V198replaced with A, G, I, L, S, T, or M; H199 replaced with K, or R; V200replaced with A, G, I, L, S, T, or M; F201 replaced with W, or Y; G202replaced with A, I, L, S, T, M, or V; D203 replaced with E; E204replaced with D; L205 replaced with A, G, I, S, T, M, or V; S206replaced with A, G, I, L, T, M, or V; L207 replaced with A, G, I, S, T,M, or V; V208 replaced with A, G, I, L, S, T, or M; T209 replaced withA, G, I, L, S, M, or V; L210 replaced with A, G, I, S, T, M, or V; F211replaced with W, or Y; R212 replaced with H, or K; I214 replaced with A,G, L, S, T, M, or V; Q215 replaced with N; N216 replaced with Q; M217replaced with A, G, I, L, S, T, or V; E219 replaced with D; T220replaced with A, G, I, L, S, M, or V; L221 replaced with A, G, I, S, T,M, or V; N223 replaced with Q; N224 replaced with Q; S225 replaced withA, G, I, L, T, M, or V; Y227 replaced with F, or W; S228 replaced withA, G, I, L, T, M, or V; A229 replaced with G, I, L, S, T, M, or V; G230replaced with A, I, L, S, T, M, or V; I231 replaced with A, G, L, S, T,M, or V; A232 replaced with G, I, L, S, T, M, or V; K233 replaced withH, or R; L234 replaced with A, G, I, S, T, M, or V; E235 replaced withD; E236 replaced with D; G237 replaced with A, I, L, S, T, M, or V; D238replaced with E; E239 replaced with D; L240 replaced with A, G, I, S, T,M, or V; Q241 replaced with N; L242 replaced with A, G, I, S, T, M, orV; A243 replaced with G, I, L, S, T, M, or V; I244 replaced with A, G,L, S, T, M, or V; R246 replaced with H, or K; E247 replaced with D; N248replaced with Q; A249 replaced with G, I, L, S, T, M, or V; Q250replaced with N; I251 replaced with A, G, L, S, T, M, or V; S252replaced with A, G, I, L, T, M, or V; L253 replaced with A, G, I, S, T,M, or V; D254 replaced with E; G255 replaced with A, I, L, S, T, M, orV; D256 replaced with E; V257 replaced with A, G, I, L, S, T, or M; T258replaced with A, G, I, L, S, M, or V; F259 replaced with W, or Y; F260replaced with W, or Y; G261 replaced with A, I, L, S, T, M, or V; A262replaced with G, I, L, S, T, M, or V; L263 replaced with A, G, I, S, T,M, or V; K264 replaced with H, or R; L265 replaced with A, G, I, S, T,M, or V; and/or L266 replaced with A, G, I, S, T, M, or V.

In another embodiment, site directed changes at the amino acid level ofB Lymphocyte Stimulator can be made by replacing a particular amino acidwith a conservative substitution. Antibodies of the present inventionmay bind B Lymphocyte Stimulator amino acid sequences containingconservative substitution mutations of the polypeptide of any one of SEQID NOS:3230-3237.

Amino acids in the B Lymphocyte Stimulator polypeptides that areessential for function can be identified by methods known in the art,such as site-directed mutagenesis or alanine-scanning mutagenesis(Cunningham and Wells, Science 244:1081-1085 (1989)). The latterprocedure introduces single alanine mutations at every residue in themolecule. The resulting mutant molecules are then tested for functionalactivity, such ligand binding and the ability to stimulate lymphocyte(e.g., B cell) as, for example, proliferation, differentiation, and/oractivation. Accordingly, antibodies of the present invention may bindamino acids in the B Lymphocyte Stimulator polypeptides that areessential for function. In preferred embodiments, antibodies of thepresent invention bind amino acids in the B Lymphocyte Stimulatorpolypeptides that are essential for function and inhibit B LymphocyteStimulator polypeptide function. In other preferred embodiments,antibodies of the present invention bind amino acids in the B LymphocyteStimulator polypeptides that are essential for function and enhance BLymphocyte Stimulator polypeptide function.

Of special interest are substitutions of charged amino acids with othercharged or neutral amino acids which may produce proteins with highlydesirable improved characteristics, such as less aggregation.Aggregation may not only reduce activity but also be problematic whenpreparing pharmaceutical formulations, because aggregates can beimmunogenic (Pinckard et al., Clin. Exp. Immunol. 2:331-340 (1967);Robbins et al., Diabetes 36: 838-845 (1987); Cleland et al., Crit. Rev.Therapeutic Drug Carrier Systems 10:307-377 (1993).

In another embodiment, the invention provides for antibodies that bindpolypeptides having amino acid sequences containing non-conservativesubstitutions of the amino acid sequence provided in SEQ ID NO:3228. Forexample, non-conservative substitutions of the B Lymphocyte Stimulatorprotein sequence provided in SEQ ID NO:3228 include: M1 replaced with D,E, H, K, R, N, Q, F, W, Y, P, or C; D2 replaced with H, K, R, A, G, I,L, S, T, M, V, N, Q, F, W, Y, P, or C; D3 replaced with H, K, R, A, G,I, L, S, T, M, V, N, Q, F, W, Y, P, or C; S4 replaced with D, E, H, K,R, N, Q, F, W, Y, P, or C; T5 replaced with D, E, H, K, R, N, Q, F, W,Y, P, or C; E6 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F,W, Y, P, or C; R7 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F,W, Y, P, or C; E8 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; Q9 replaced with D, E, H, K, R, A, G, I, L, S, T, M,V, F, W, Y, P, or C; S10 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; R11 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; L12 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; T13replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S14 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; C15 replaced with D, E, H, K, R,A, G, I, L, S, T, M, V, N, Q, F, W, Y, or P; L16 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; K17 replaced with D, E, A, G, I, L, S, T,M, V, N, Q, F, W, Y, P, or C; K18 replaced with D, E, A, G, I, L, S, T,M, V, N, Q, F, W, Y, P, or C; R19 replaced with D, E, A, G, I, L, S, T,M, V, N, Q, F, W, Y, P, or C; E20 replaced with H, K, R, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; E21 replaced with H, K, R, A, G, I, L,S, T, M, V, N, Q, F, W, Y, P, or C; M22 replaced with D, E, H, K, R, N,Q, F, W, Y, P, or C; K23 replaced with D, E, A, G, I, L, S, T, M, V, N,Q, F, W, Y, P, or C; L24 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; K25 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; E26 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; C27 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, or P; V28 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;S29 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; I30 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; L31 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; P32 replaced with D, E, H, K, R, A, G, I,L, S, T, M, V, N, Q, F, W, Y, or C; R33 replaced with D, E, A, G, I, L,S, T, M, V, N, Q, F, W, Y, P, or C; K34 replaced with D, E, A, G, I, L,S, T, M, V, N, Q, F, W, Y, P, or C; E35 replaced with H, K, R, A, G, I,L, S, T, M, V, N, Q, F, W, Y, P, or C; S36 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; P37 replaced with D, E, H, K, R, A, G, I, L, S,T, M, V, N, Q, F, W, Y, or C; S38 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; V39 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;R40 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;S41 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S42 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; K43 replaced with D, E, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; D44 replaced with H, K, R,A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; G45 replaced with D, E,H, K, R, N, Q, F, W, Y, P, or C; K46 replaced with D, E, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; L47 replaced with D, E, H, K, R, N, Q,F, W, Y, P, or C; L48 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; A49 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A50 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; T51 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; L52 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; L53 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;L54 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A55 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; L56 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; L57 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; S58 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;C59 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y,or P; C60 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F,W, Y, or P; L61 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; T62replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; V63 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; V64 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; S65 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; F66 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V,P, or C; Y67 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V,P, or C; Q68 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W,Y, P, or C; V69 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A70replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A71 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; L72 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; Q73 replaced with D, E, H, K, R, A, G, I, L, S,T, M, V, F, W, Y, P, or C; G74 replaced with D, E, H, K, R, N, Q, F, W,Y, P, or C; D75 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F,W, Y, P, or C; L76 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;A77 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S78 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; L79 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; R80 replaced with D, E, A, G, I, L, S, T,M, V, N, Q, F, W, Y, P, or C; A81 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; E82 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; L83 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; Q84 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P,or C; G85 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; H86replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; H87replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; A88replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; E89 replaced withH, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; K90 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L91 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; P92 replaced with D, E, H,K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; A93 replaced with D,E, H, K, R, N, Q, F, W, Y, P, or C; G94 replaced with D, E, H, K, R, N,Q, F, W, Y, P, or C; A95 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; G96 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A97replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P98 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; K99 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; A100 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; G101 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; L102 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; E103 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; E104 replaced with H, K, R, A, G, I, L, S, T, M, V, N,Q, F, W, Y, P, or C; A105 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; P106 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F,W, Y, or C; A107 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;V108 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; T109 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; A110 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; G111 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; L112 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;K113 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;I114 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; F115 replacedwith D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; E116 replacedwith H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; P117replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; P118 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W,Y, or C; A119 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P120replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; G121 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; E122replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;G123 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; N124 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; S125replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S126 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; Q127 replaced with D, E, H, K, R,A, G, I, L, S, T, M, V, F, W, Y, P, or C; N128 replaced with D, E, H, K,R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; S129 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; R130 replaced with D, E, A, G, I, L, S, T,M, V, N, Q, F, W, Y, P, or C; N131 replaced with D, E, H, K, R, A, G, I,L, S, T, M, V, F, W, Y, P, or C; K132 replaced with D, E, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; R133 replaced with D, E, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; A134 replaced with D, E, H, K, R, N, Q,F, W, Y, P, or C; V135 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; Q136 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P,or C; G137 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P138replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; E139 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; E140 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; T141 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; V142replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; T143 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; Q144 replaced with D, E, H, K, R,A, G, I, L, S, T, M, V, F, W, Y, P, or C; D145 replaced with H, K, R, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; C146 replaced with D, E, H,K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or P; L147 replaced with D,E, H, K, R, N, Q, F, W, Y, P, or C; Q148 replaced with D, E, H, K, R, A,G, I, L, S, T, M, V, F, W, Y, P, or C; L149 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; I150 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; A151 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; D152replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;S153 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; E154 replacedwith H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; T155replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P156 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; T157replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; I158 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; Q159 replaced with D, E, H, K, R,A, G, I, L, S, T, M, V, F, W, Y, P, or C; K160 replaced with D, E, A, G,I, L, S, T, M, V, N, Q, F, W, Y, P, or C; G161 replaced with D, E, H, K,R, N, Q, F, W, Y, P, or C; S162 replaced with D, E, H, K, R, N, Q, F, W,Y, P, or C; Y163 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M,V, P, or C; T164 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;F165 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C;V166 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P167 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; W168replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; L169replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L170 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; S171 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; F172 replaced with D, E, H, K, R, N, Q, A, G, I,L, S, T, M, V, P, or C; K173 replaced with D, E, A, G, I, L, S, T, M, V,N, Q, F, W, Y, P, or C; R174 replaced with D, E, A, G, I, L, S, T, M, V,N, Q, F, W, Y, P, or C; G175 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; S176 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A177replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L178 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; E179 replaced with H, K, R, A, G,I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E180 replaced with H, K, R, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; K181 replaced with D, E, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E182 replaced with H, K, R,A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; N183 replaced with D, E,H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; K184 replaced with D,E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; I185 replaced with D,E, H, K, R, N, Q, F, W, Y, P, or C; L186 replaced with D, E, H, K, R, N,Q, F, W, Y, P, or C; V187 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; K188 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; E189 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; T190 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; G191replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Y192 replaced withD, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; F193 replaced withD, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; F194 replaced withD, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; I195 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; Y196 replaced with D, E, H, K, R,N, Q, A, G, I, L, S, T, M, V, P, or C; G197 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; Q198 replaced with D, E, H, K, R, A, G, I, L, S,T, M, V, F, W, Y, P, or C; V199 replaced with D, E, H, K, R, N, Q, F, W,Y, P, or C; L200 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;Y201 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C;T202 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; D203 replacedwith H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; K204replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; T205replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Y206 replaced withD, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; A207 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; M208 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; G209 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; H210 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; L211 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; I212replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Q213 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; R214 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; K215 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; K216 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; V217 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; H218 replaced with D, E, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; V219 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; F220 replaced with D, E, H, K, R, N, Q, A,G, I, L, S, T, M, V, P, or C; G221 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; D222 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; E223 replaced with H, K, R, A, G, I, L, S, T, M, V, N,Q, F, W, Y, P, or C; L224 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; S225 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L226replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; V227 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; T228 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; L229 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; F230 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V,P, or C; R231 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; C232 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, or P; I233 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;Q234 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, orC; N235 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P,or C; M236 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P237replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; E238 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; T239 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L240replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P241 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; N242replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C;N243 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, orC; S244 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; C245replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orP; Y246 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, orC; S247 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A248replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; G249 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; I250 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; A251 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; K252 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; L253 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; E254replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;E255 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, orC; G256 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; D257replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;E258 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, orC; L259 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Q260replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C;L261 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A262 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; I263 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; P264 replaced with D, E, H, K, R, A, G, I,L, S, T, M, V, N, Q, F, W, Y, or C; R265 replaced with D, E, A, G, I, L,S, T, M, V, N, Q, F, W, Y, P, or C; E266 replaced with H, K, R, A, G, I,L, S, T, M, V, N, Q, F, W, Y, P, or C; N267 replaced with D, E, H, K, R,A, G, I, L, S, T, M, V, F, W, Y, P, or C; A268 replaced with D, E, H, K,R, N, Q, F, W, Y, P, or C; Q269 replaced with D, E, H, K, R, A, G, I, L,S, T, M, V, F, W, Y, P, or C; I270 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; S271 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;L272 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; D273 replacedwith H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; G274replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; D275 replaced withH, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; V276 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; T277 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; F278 replaced with D, E, H, K, R, N, Q, A,G, I, L, S, T, M, V, P, or C; F279 replaced with D, E, H, K, R, N, Q, A,G, I, L, S, T, M, V, P, or C; G280 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; A281 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;L282 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; K283 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L284 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; and/or L285 replaced with D,E, H, K, R, N, Q, F, W, Y, P, or C.

In an additional embodiment, antibodies of the present invention bind BLymphocyte Stimulator polypeptides comprising, or alternativelyconsisting of, a B Lymphocyte Stimulator amino acid sequence in whichmore than one amino acid (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30and 50) is replaced with the substituted amino acids as described above(either conservative or nonconservative).

In another embodiment of the invention, antibodies of the presentinvention bind B Lymphocyte Stimulator polypeptides withnon-conservative substitutions of the sequence provided in SEQ IDNO:3229 including: M1 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; D2 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; D3 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; S4 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; T5replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; E6 replaced with H,K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; R7 replaced withD, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E8 replaced withH, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; Q9 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; S10replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; R11 replaced withD, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L12 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; T13 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; S14 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; C15 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, or P; L16 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;K17 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;K18 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;R19 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;E20 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, orC; E21 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; M22 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; K23replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L24replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; K25 replaced withD, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E26 replaced withH, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; C27 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or P; V28replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S29 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; I30 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; L31 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; P32 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, or C; R33 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F,W, Y, P, or C; K34 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F,W, Y, P, or C; E35 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; S36 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; P37 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W,Y, or C; S38 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; V39replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; R40 replaced withD, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; S41 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; S42 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; K43 replaced with D, E, A, G, I, L, S, T, M, V,N, Q, F, W, Y, P, or C; D44 replaced with H, K, R, A, G, I, L, S, T, M,V, N, Q, F, W, Y, P, or C; G45 replaced with D, E, H, K, R, N, Q, F, W,Y, P, or C; K46 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W,Y, P, or C; L47 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L48replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A49 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; A50 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; T51 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; L52 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L53replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L54 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; A55 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; L56 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; L57 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S58replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; C59 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or P; C60 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or P; L61replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; T62 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; V63 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; V64 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; S65 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; F66replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; Y67replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; Q68replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C;V69 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A70 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; A71 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; L72 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; Q73 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V,F, W, Y, P, or C; G74 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; D75 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; L76 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A77replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S78 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; L79 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; R80 replaced with D, E, A, G, I, L, S, T, M, V,N, Q, F, W, Y, P, or C; A81 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; E82 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W,Y, P, or C; L83 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Q84replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C;G85 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; H86 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; H87 replacedwith D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; A88 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; E89 replaced with H, K, R,A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; K90 replaced with D, E,A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L91 replaced with D, E,H, K, R, N, Q, F, W, Y, P, or C; P92 replaced with D, E, H, K, R, A, G,I, L, S, T, M, V, N, Q, F, W, Y, or C; A93 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; G94 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; A95 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; G96replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A97 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; P98 replaced with D, E, H, K, R,A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; K99 replaced with D, E, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; A100 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; G101 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; L102 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;E103 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, orC; E104 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; A105 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P106replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; A107 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; V108replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; T109 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; A110 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; G111 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; L112 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; K113replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; I114replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; F115 replaced withD, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; E116 replaced withH, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; P117 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; P118replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; A119 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P120replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; G121 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; E122replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;G123 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; N124 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; S125replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S126 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; Q127 replaced with D, E, H, K, R,A, G, I, L, S, T, M, V, F, W, Y, P, or C; N128 replaced with D, E, H, K,R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; S129 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; R130 replaced with D, E, A, G, I, L, S, T,M, V, N, Q, F, W, Y, P, or C; N131 replaced with D, E, H, K, R, A, G, I,L, S, T, M, V, F, W, Y, P, or C; K132 replaced with D, E, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; R133 replaced with D, E, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; A134 replaced with D, E, H, K, R, N, Q,F, W, Y, P, or C; V135 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; Q136 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P,or C; G137 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P138replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, orC; E139 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; E140 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y,P, or C; T141 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; G142replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S143 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; Y144 replaced with D, E, H, K, R,N, Q, A, G, I, L, S, T, M, V, P, or C; T145 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; F146 replaced with D, E, H, K, R, N, Q, A, G, I,L, S, T, M, V, P, or C; V147 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; P148 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, N, Q,F, W, Y, or C; W149 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T,M, V, P, or C; L150 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;L151 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S152 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; F153 replaced with D, E, H,K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; K154 replaced with D, E, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; R155 replaced with D, E, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; G156 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; S157 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; A158 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;L159 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; E160 replacedwith H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E161replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;K162 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;E163 replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, orC; N164 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P,or C; K165 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P,or C; I166 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L167replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; V168 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; K169 replaced with D, E, A, G, I,L, S, T, M, V, N, Q, F, W, Y, P, or C; E170 replaced with H, K, R, A, G,I, L, S, T, M, V, N, Q, F, W, Y, P, or C; T171 replaced with D, E, H, K,R, N, Q, F, W, Y, P, or C; G172 replaced with D, E, H, K, R, N, Q, F, W,Y, P, or C; Y173 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M,V, P, or C; F174 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M,V, P, or C; F175 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M,V, P, or C; I176 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;Y177 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C;G178 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Q179 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; V180replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L181 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; Y182 replaced with D, E, H, K, R,N, Q, A, G, I, L, S, T, M, V, P, or C; T183 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; D184 replaced with H, K, R, A, G, I, L, S, T, M,V, N, Q, F, W, Y, P, or C; K185 replaced with D, E, A, G, I, L, S, T, M,V, N, Q, F, W, Y, P, or C; T186 replaced with D, E, H, K, R, N, Q, F, W,Y, P, or C; Y187 replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M,V, P, or C; A188 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C;M189 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; G190 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; H191 replaced with D, E, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L192 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; I193 replaced with D, E, H, K, R, N, Q, F,W, Y, P, or C; Q194 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V,F, W, Y, P, or C; R195 replaced with D, E, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; K196 replaced with D, E, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; K197 replaced with D, E, A, G, I, L, S, T, M, V, N, Q,F, W, Y, P, or C; V198 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC; H199 replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, orC; V200 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; F201replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; G202replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; D203 replaced withH, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E204 replacedwith H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L205replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S206 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; L207 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; V208 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; T209 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L210replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; F211 replaced withD, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; R212 replaced withD, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; C213 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or P; I214replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Q215 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; N216 replacedwith D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; M217replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P218 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; E219replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;T220 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; L221 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; P222 replaced with D, E, H,K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; N223 replaced with D,E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; N224 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C; S225 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; C226 replaced with D, E, H,K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or P; Y227 replaced with D,E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; S228 replaced with D,E, H, K, R, N, Q, F, W, Y, P, or C; A229 replaced with D, E, H, K, R, N,Q, F, W, Y, P, or C; G230 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; I231 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A232replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; K233 replaced withD, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; L234 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; E235 replaced with H, K, R, A, G,I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E236 replaced with H, K, R, A,G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; G237 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; D238 replaced with H, K, R, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; E239 replaced with H, K, R, A, G, I, L,S, T, M, V, N, Q, F, W, Y, P, or C; L240 replaced with D, E, H, K, R, N,Q, F, W, Y, P, or C; Q241 replaced with D, E, H, K, R, A, G, I, L, S, T,M, V, F, W, Y, P, or C; L242 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; A243 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; I244replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; P245 replaced withD, E, H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, or C; R246replaced with D, E, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C; E247replaced with H, K, R, A, G, I, L, S, T, M, V, N, Q, F, W, Y, P, or C;N248 replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, orC; A249 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; Q250replaced with D, E, H, K, R, A, G, I, L, S, T, M, V, F, W, Y, P, or C;I251 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; S252 replacedwith D, E, H, K, R, N, Q, F, W, Y, P, or C; L253 replaced with D, E, H,K, R, N, Q, F, W, Y, P, or C; D254 replaced with H, K, R, A, G, I, L, S,T, M, V, N, Q, F, W, Y, P, or C; G255 replaced with D, E, H, K, R, N, Q,F, W, Y, P, or C; D256 replaced with H, K, R, A, G, I, L, S, T, M, V, N,Q, F, W, Y, P, or C; V257 replaced with D, E, H, K, R, N, Q, F, W, Y, P,or C; T258 replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; F259replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; F260replaced with D, E, H, K, R, N, Q, A, G, I, L, S, T, M, V, P, or C; G261replaced with D, E, H, K, R, N, Q, F, W, Y, P, or C; A262 replaced withD, E, H, K, R, N, Q, F, W, Y, P, or C; L263 replaced with D, E, H, K, R,N, Q, F, W, Y, P, or C; K264 replaced with D, E, A, G, I, L, S, T, M, V,N, Q, F, W, Y, P, or C; L265 replaced with D, E, H, K, R, N, Q, F, W, Y,P, or C; and/or L266 replaced with D, E, H, K, R, N, Q, F, W, Y, P, orC.

In another embodiment, site directed changes at the amino acid level ofB Lymphocyte Stimulator can be made by replacing a particular amino acidwith a non-conservative substitution. Antibodies of the presentinvention may bind B Lymphocyte Stimulator amino acid sequencescontaining non-conservative substitution mutations of the polypeptide ofany one of SEQ ID NOS:3230-3237.

In an additional embodiment, antibodies of the present invention bind BLymphocyte Stimulator polypeptides comprising, or alternativelyconsisting of, a B Lymphocyte Stimulator amino acid sequence in whichmore than one amino acid (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 30and 50) is replaced with the substituted amino acids as described above(either conservative or nonconservative).

Replacement of amino acids can also change the selectivity of thebinding of a ligand to cell surface receptors. For example, Ostade etal., Nature 361:266-268 (1993) describes certain mutations resulting inselective binding of TNF-alpha to only one of the two known types of TNFreceptors. Since B Lymphocyte Stimulator is a member of the TNFpolypeptide family, mutations similar to those in TNF-alpha are likelyto have similar effects in B Lymphocyte Stimulator polypeptides.

Sites that are critical for ligand-receptor binding can also bedetermined by structural analysis such as crystallization, nuclearmagnetic resonance or photoaffinity labeling (Smith et al., J. Mol.Biol. 224:899-904 (1992) and de Vos et al. Science 255:306-312 (1992)).

Since B Lymphocyte Stimulator is a member of the TNF-related proteinfamily, mutations may be made in sequences encoding amino acids in theTNF conserved domain, e.g., in positions Gly-191 through Leu-284 of SEQID NO:3228 or in positions Gly-172 through Leu-265 of SEQ ID NO:3229,may modulate rather than completely eliminate functional activities(e.g., biological activities) of B Lymphocyte Stimulator polypeptides orfragments or variants thereof. Accordingly, antibodies of the presentinvention may bind B Lymphocyte Stimulator polypeptides that havemutations in the TNF conserved domain. In preferred embodiments,antibodies of the present invention may bind B Lymphocyte Stimulatorpolypeptides that have mutations in the TNF conserved domain and act asantagonists of B Lymphocyte Stimulator. In other preferred embodiments,antibodies of the present invention may bind B Lymphocyte Stimulatorpolypeptides that have mutations in the TNF conserved domain and act asagonists of B Lymphocyte Stimulator.

Recombinant DNA technology known to those skilled in the art (see, forinstance, DNA shuffling supra) can be used to create novel mutantproteins or muteins including single or multiple amino acidsubstitutions, deletions, additions or fusion proteins. Such modifiedpolypeptides can show, e g , enhanced activity or increased stability.In addition, they may be purified in higher yields and show bettersolubility than the corresponding natural polypeptide, at least undercertain purification and storage conditions.

Thus, the invention also encompasses antibodies that bind B LymphocyteStimulator derivatives and analogs that have one or more amino acidresidues deleted, added, or substituted to generate B LymphocyteStimulator polypeptides, e.g., that are better suited for expression,scale up, etc., in the host cells. For example, cysteine residues can bedeleted or substituted with another amino acid residue in order toeliminate disulfide bridges; N-linked glycosylation sites can be alteredor eliminated to achieve, for example, expression of a homogeneousproduct that is more easily recovered and purified from yeast hostswhich are known to hyperglycosylate N-linked sites. To this end, avariety of amino acid substitutions at one or both of the first or thirdamino acid positions on any one or more of the glycosylation recognitionsequences in the B Lymphocyte Stimulator polypeptides of the invention,and/or an amino acid deletion at the second position of any one or moresuch recognition sequences will prevent glycosylation of the BLymphocyte Stimulator at the modified tripeptide sequence (see, e.g.,Miyajimo et al., EMBO J 5(6):1193-1197). By way of non-limiting example,mutation of the serine at position 244 to alanine either singly or incombination with mutation of the asparagine at position 242 to glutamineabolishes glycosylation of the mature soluble form of B LymphocyteStimulator (e.g., amino acids 134-285 of SEQ ID NO:3228) when expressedin the yeast Pichea pastoris. A mutant B Lymphocyte Stimulatorpolypeptide in which only the asparagine at position 242 is mutated toglutamine, is still gycosylated when expressed in Pichea pastoris. Inthis mutant, the glycosylation event may be due to the activation orunmasking of an O-linked glyscosylation site at serine 244. Similarmutations affecting glycosylation could also be made in the B LymphocyteStimulator polypeptide of SEQ ID NO:3229, i.e., aspargine-223 toglutamine and/or serine-224 to alanine of SEQ ID NO:3229. Additionally,one or more of the amino acid residues of the polypeptides of theinvention (e.g., arginine and lysine residues) may be deleted orsubstituted with another residue to eliminate undesired processing byproteases such as, for example, furins or kexins. One possible result ofsuch a mutation is that B Lymphocyte Stimulator polypeptide of theinvention is not cleaved and released from the cell surface.Accordingly, antibodies of the invention may bind B LymphocyteStimulator derivatives and analogs that have one or more amino acidresidues deleted, added, or substituted. In other embodiments,antibodies of the invention may bind B Lymphocyte Stimulatorderivatives, variants or analogs that are unable to be cleaved from thecell surface.

In a specific embodiment, antibodies of the invention bind B LymphocyteStimulator polypeptides in which Lys-132 and/or Arg-133 of the BLymphocyte Stimulator sequence shown in SEQ ID NO:3228 is mutated toanother amino acid residue, or deleted altogether, to prevent ordiminish release of the soluble form of B Lymphocyte Stimulator fromcells expressing B Lymphocyte Stimulator. In a more specific embodiment,antibodies of the invention bind B Lymphocyte Stimulator polypeptides inwhich Lys-132 of the B Lymphocyte Stimulator sequence shown in SEQ IDNO:3228 is mutated to Ala-132. In another, nonexclusive specificembodiment, antibodies of the invention bind B Lymphocyte Stimulatorpolypeptides in which Arg-133 of the B Lymphocyte Stimulator sequenceshown in SEQ ID NO:3228 is mutated to Ala-133. These mutated proteinshave uses such as, for example, in ex vivo therapy or gene therapy, toengineer cells expressing a B Lymphocyte Stimulator polypeptide that isretained on the surface of the engineered cells.

In a specific embodiment, antibodies of the invention bind B LymphocyteStimulator polypeptides in which Cys-146 of the B Lymphocyte Stimulatorsequence shown in SEQ ID NO:3228 is mutated to another amino acidresidue, or deleted altogether, for example, to aid preventing ordiminishing oligomerization of the mutant B Lymphocyte Stimulatorpolypeptide when expressed in an expression system. In a specificembodiment, antibodies of the invention bind B Lymphocyte Stimulatorpolypeptides in which Cys-146 is replaced with a serine amino acidresidue.

In another specific embodiment, antibodies of the invention bind BLymphocyte Stimulator polypeptides in which Cys-232 of the B LymphocyteStimulator sequence shown in SEQ ID NO:3228 is mutated to another aminoacid residue, or deleted altogether, for example, to aid preventing ordiminishing oligomerization of the mutant B Lymphocyte Stimulatorpolypeptide when expressed in an expression system. In a specificembodiment, antibodies of the invention bind B Lymphocyte Stimulatorpolypeptides in which Cys-232 is replaced with a serine amino acidresidue. Polypeptides encoding these polypeptides are also encompassedby the invention.

In yet another specific embodiment, antibodies of the invention bind BLymphocyte Stimulator polypeptides in which Cys-245 of the B LymphocyteStimulator sequence shown in SEQ ID NO:3228 is mutated to another aminoacid residue, or deleted altogether, for example, to aid preventing ordiminishing oligomerization of the mutant B Lymphocyte Stimulatorpolypeptide when expressed in an expression system. In a specificembodiment, antibodies of the invention bind B Lymphocyte Stimulatorpolypeptides in which Cys-245 is replaced with a serine amino acidresidue. Polypeptides encoding these polypeptides are also encompassedby the invention.

The polypeptides of the present invention are preferably provided in anisolated form, and preferably are substantially purified. Arecombinantly produced version of the B Lymphocyte Stimulatorpolypeptides can be substantially purified by the one-step methoddescribed in Smith and Johnson, Gene 67:31-40 (1988).

The antibodies of the present invention bind B Lymphocyte Stimulatorpolypeptides including the complete polypeptide encoded by the depositedcDNA (ATCC™ Deposit No. 97768) including the intracellular,transmembrane and extracellular domains of the polypeptide encoded bythe deposited cDNA, the mature soluble polypeptide encoded by thedeposited cDNA, the extracellular domain minus the intracellular andtransmembrane domains of the protein, the complete polypeptide of SEQ IDNO:3228, the mature soluble polypeptide of SEQ ID NO:3228, e.g., aminoacids 134-285 of SEQ ID NO:3228, the extracellular domain of SEQ IDNO:3228, amino acid residues 73-285 of SEQ ID NO:3228 minus theintracellular and transmembrane domains, as well as polypeptides whichhave at least 80%, 85%, 90% similarity, more preferably at least 95%similarity, and still more preferably at least 96%, 97%, 98% or 99%similarity to those described above. Polynucleotides encoding thesepolypeptides are also encompassed by the invention.

The antibodies of the present invention bind B Lymphocyte Stimulatorpolypeptides including the complete polypeptide encoded by the depositedcDNA including the intracellular, transmembrane and extracellulardomains of the polypeptide encoded by the deposited cDNA (ATCC™ DepositNo. 203518), the mature soluble polypeptide encoded by the depositedcDNA, the extracellular domain minus the intracellular and transmembranedomains of the protein, the complete polypeptide of SEQ ID NO:3229, themature soluble of SEQ ID NO:3229, e.g., amino acid residues 134-266 ofSEQ ID NO:3229, the extracellular domain of SEQ ID NO:3229, e.g., aminoacid residues 73-266 of SEQ ID NO:3229 minus the intracellular andtransmembrane domains, as well as polypeptides which have at least 80%,85%, 90% similarity, more preferably at least 95% similarity, and stillmore preferably at least 96%, 97%, 98% or 99% similarity to thosedescribed above. Polynucleotides encoding these polypeptides are alsoencompassed by the invention.

Further antibodies of the present invention bind polypeptides includingpolypeptides at least 80%, or at least 85% identical, more preferably atleast 90% or 95% identical, still more preferably at least 96%, 97%, 98%or 99% identical to the polypeptide encoded by the deposited cDNA (ATCC™Deposit No. 97768) or to the polypeptide of SEQ ID NO:3228, and alsoinclude antibodies that bind portions of such polypeptides with at least30 amino acids and more preferably at least 50 amino acids.

Further antibodies of the present invention bind polypeptides includingpolypeptides at least 80%, or at least 85% identical, more preferably atleast 90% or 95% identical, still more preferably at least 96%, 97%, 98%or 99% identical to the polypeptide encoded by the deposited cDNA (ATCC™Deposit No. 203518) or to the polypeptide of SEQ ID NO:3229, and alsoinclude antibodies that bind portions of such polypeptides with at least30 amino acids and more preferably at least 50 amino acids.Polynucleotides encoding these polypeptides are also encompassed by theinvention.

By “% similarity” for two polypeptides is intended a similarity scoreproduced by comparing the amino acid sequences of the two polypeptidesusing the Bestfit program (Wisconsin Sequence Analysis Package, Version8 for Unix, Genetics Computer Group, University Research Park, 575Science Drive, Madison, Wis. 53711) and the default settings fordetermining similarity. Bestfit uses the local homology algorithm ofSmith and Waterman (Advances in Applied Mathematics 2:482-489, 1981) tofind the best segment of similarity between two sequences.

By a polypeptide having an amino acid sequence at least, for example,95% “identical” to a reference amino acid sequence of a B LymphocyteStimulator polypeptide is intended that the amino acid sequence of thepolypeptide is identical to the reference sequence except that thepolypeptide sequence may include up to five amino acid alterations pereach 100 amino acids of the reference amino acid of the B LymphocyteStimulator polypeptide. In other words, to obtain a polypeptide havingan amino acid sequence at least 95% identical to a reference amino acidsequence, up to 5% of the amino acid residues in the reference sequencemay be deleted or substituted with another amino acid, or a number ofamino acids up to 5% of the total amino acid residues in the referencesequence may be inserted into the reference sequence. These alterationsof the reference sequence may occur at the amino or carboxy terminalpositions of the reference amino acid sequence or anywhere between thoseterminal positions, interspersed either individually among residues inthe reference sequence or in one or more contiguous groups within thereference sequence.

As a practical matter, whether any particular polypeptide is at least80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to, for instance, theamino acid sequence of SEQ ID NO:3228, the amino acid sequence encodedby the deposited cDNA clone HNEDU15 (ATCC™ Accession No. 97768), orfragments thereof, or, for instance, to the amino acid sequence of SEQID NO:3229, the amino acid sequence encoded by the deposited cDNA cloneHDPMC52 (ATCC™ Accession No. 203518), or fragments thereof, can bedetermined conventionally using known computer programs such the Bestfitprogram (Wisconsin Sequence Analysis Package, Version 8 for Unix,Genetics Computer Group, University Research Park, 575 Science Drive,Madison, Wis. 53711). When using Bestfit or any other sequence alignmentprogram to determine whether a particular sequence is, for instance, 95%identical to a reference sequence according to the present invention,the parameters are set, of course, such that the percentage of identityis calculated over the full length of the reference amino acid sequenceand that gaps in homology of up to 5% of the total number of amino acidresidues in the reference sequence are allowed.

In a specific embodiment, the identity between a reference (query)sequence (a sequence of the present invention) and a subject sequence,also referred to as a global sequence alignment, is determined using theFASTDB computer program based on the algorithm of Brutlag et al. (Comp.App. Biosci. 6:237-245 (1990)). Preferred parameters used in a FASTDBamino acid alignment are: Matrix=PAM 0, k-tuple=2, Mismatch Penalty=1,Joining Penalty=20, Randomization Group Length=0, Cutoff Score=1, WindowSize=sequence length, Gap Penalty=5, Gap Size Penalty=0.05, WindowSize=500 or the length of the subject amino acid sequence, whichever isshorter. According to this embodiment, if the subject sequence isshorter than the query sequence due to N- or C-terminal deletions, notbecause of internal deletions, a manual correction is made to theresults to take into consideration the fact that the FASTDB program doesnot account for N- and C-terminal truncations of the subject sequencewhen calculating global percent identity. For subject sequencestruncated at the N- and C-termini, relative to the query sequence, thepercent identity is corrected by calculating the number of residues ofthe query sequence that are N- and C-terminal of the subject sequence,which are not matched/aligned with a corresponding subject residue, as apercent of the total bases of the query sequence. A determination ofwhether a residue is matched/aligned is determined by results of theFASTDB sequence alignment. This percentage is then subtracted from thepercent identity, calculated by the above FASTDB program using thespecified parameters, to arrive at a final percent identity score. Thisfinal percent identity score is what is used for the purposes of thisembodiment. Only residues to the N- and C-termini of the subjectsequence, which are not matched/aligned with the query sequence, areconsidered for the purposes of manually adjusting the percent identityscore. That is, only query residue positions outside the farthest N- andC-terminal residues of the subject sequence. For example, a 90 aminoacid residue subject sequence is aligned with a 100 residue querysequence to determine percent identity. The deletion occurs at theN-terminus of the subject sequence and therefore, the FASTDB alignmentdoes not show a matching/alignment of the first 10 residues at theN-terminus. The 10 unpaired residues represent 10% of the sequence(number of residues at the N- and C-termini not matched/total number ofresidues in the query sequence) so 10% is subtracted from the percentidentity score calculated by the FASTDB program. If the remaining 90residues were perfectly matched the final percent identity would be 90%.In another example, a 90 residue subject sequence is compared with a 100residue query sequence. This time the deletions are internal deletionsso there are no residues at the N- or C-termini of the subject sequencewhich are not matched/aligned with the query. In this case the percentidentity calculated by FASTDB is not manually corrected. Once again,only residue positions outside the N- and C-terminal ends of the subjectsequence, as displayed in the FASTDB alignment, which are notmatched/aligned with the query sequence are manually corrected for. Noother manual corrections are made for the purposes of this embodiment.

Antibodies that Immunospecifically Bind B Lymphocyte StimulatorPolypeptides

The present invention also encompasses antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to B LymphocyteStimulator polypeptides, which antibodies comprise, or alternativelyconsist of, all or a portion of a heavy and/or light chain variabledomain of the scFvs referred to in Table 1.

The present invention also encompasses methods and compositions fordetecting, diagnosing and/or prognosing diseases or disorders associatedwith aberrant B Lymphocyte Stimulator or B Lymphocyte Stimulatorreceptor expression or inappropriate B Lymphocyte Stimulator or BLymphocyte Stimulator receptor function in an animal, preferably amammal, and most preferably a human, comprising using antibodies(including molecules which comprise, or alternatively consist of,antibody fragments or variants thereof) that immunospecifically bind toB Lymphocyte Stimulator. Diseases and disorders which can be detected,diagnosed or prognosed with the antibodies of the invention include, butare not limited to, immune disorders (e.g., lupus, rheumatoid arthritis,multiple sclerosis, myasthenia gravis, Hashimoto's disease, andimmunodeficiency syndrome), inflammatory disorders (e.g., asthma,allergic disorders, and rheumatoid arthritis), infectious diseases(e.g., AIDS), and proliferative disorders (e.g., leukemia, carcinoma,and lymphoma).

The present invention further encompasses methods and compositions forpreventing, treating or ameliorating diseases or disorders associatedwith aberrant B Lymphocyte Stimulator or B Lymphocyte Stimulatorreceptor expression or inappropriate B Lymphocyte Stimulator or BLymphocyte Stimulator receptor function in an animal, preferably amammal, and most preferably a human, comprising administering to saidanimal an effective amount of one or more antibodies (includingmolecules which comprise, or alternatively consist of, antibodyfragments or variants thereof) that immunospecifically bind to BLymphocyte Stimulator. Diseases and disorders which can be prevented,treated or inhibited by administering an effective amount of one or moreantibodies or molecules of the invention include, but are not limitedto, immune disorders (e.g., lupus, rheumatoid arthritis, multiplesclerosis, myasthenia gravis, Hashimoto's disease, and immunodeficiencysyndrome), inflammatory disorders (e.g., asthma, allergic disorders, andrheumatoid arthritis), infectious diseases (e.g., AIDS), andproliferative disorders (e.g., leukemia, carcinoma, and lymphoma).

Anti-B Lymphocyte Stimulator Antibodies

The antibodies of the present invention were discovered, in part, usingphage display technology. Single chain antibody molecules (“scFvs”)displayed on the surface of phage particles were screened to identifythose scFvs that immunospecifically bind to B Lymphocyte Stimulator,including the membrane-bound form and soluble form of B LymphocyteStimulator. The present invention encompasses the scFvs and portionsthereof that were identified to immunospecifically bind to B LymphocyteStimulator, including scFvs that immunospecifically bind to the solubleform of B Lymphocyte Stimulator, scFvs that immunospecifically bind tothe membrane-bound form of B Lymphocyte Stimulator, and scFvs thatimmunospecifically bind to both the soluble form and membrane-bound formof B Lymphocyte Stimulator. In particular, the present inventionencompasses scFvs comprising, or alternatively consisting of, the aminoacid sequence of SEQ ID NOS: 1-2128, as referred to in Table 1.Preferably, the scFvs of the present invention comprise, oralternatively consist of, the amino acid sequence of SEQ ID NOS:1-46,321-329, 834-872, 1563-1595, or 1881-1908. The scFvs include scFvs thatbind to soluble B Lymphocyte Stimulator (e.g., scFvs comprising, oralternatively consisting of, an amino acid sequence of SEQ ID NOS:1563-1880), scFvs that bind to the membrane-bound form of B LymphocyteStimulator (e.g., scFvs comprising, or alternatively consisting of, anamino acid sequence of SEQ ID NOS: 1881-2128), and scFvs that bind toboth the soluble form and the membrane-bound form of B LymphocyteStimulator (e.g., scFvs comprising, or alternatively consisting of, anamino acid sequence of SEQ ID NOS: 1-1562). Molecules comprising, oralternatively consisting of, fragments or variants of these scFvs, thatimmunospecifically bind to B Lymphocyte Stimulator are also encompassedby the invention, as are nucleic acid molecules encoding these scFvs,molecules, fragments and/or variants.

In one embodiment of the present invention, scFvs thatimmunospecifically bind to B Lymphocyte Stimulator comprise apolypeptide having the amino acid sequence of any one of the VH domainsreferred to in Table 1 and/or any one of the VL domains referred to inTable 1. In preferred embodiments, scFvs of the present inventioncomprise the amino acid sequence of a VH domain and VL domain from thesame scFv referred to in Table 1. In alternative embodiments, scFvs ofthe present invention comprise the amino acid sequence of a VH domainand VL domain from different scFvs referred to in Table 1. In anotherembodiment, scFvs that immunospecifically bind to B LymphocyteStimulator, comprise a polypeptide having the amino acid sequence of anyone, two, three, or more of the VH CDRs referred to in Table 1 and/orany one, two, three, or more of the VL CDRs referred to in Table 1. Inpreferred embodiments, scFvs of the present invention comprise the aminoacid sequence of a VH CDR and VL CDR from the same scFv referred to inTable 1. In alternative embodiments, scFvs of the present inventioncomprise the amino acid sequence of a VH CDR and VL CDR from differentscFvs referred to in Table 1. Molecules comprising, or alternativelyconsisting of, antibody fragments or variants of the scFvs referred toin Table 1 that immunospecifically bind to B Lymphocyte Stimulator arealso encompassed by the invention, as are nucleic acid moleculesencoding these scFvs, molecules, fragments and/or variants.

(Table 1 can be found at the end of the specification just prior to theclaims.)

In another embodiment of the present invention, an scFv thatimmunospecifically binds to a soluble form of B Lymphocyte Stimulator,comprises, or alternatively consists of, the amino acid sequence of SEQID NOS:1563-1880 as referred to in Table 1. In a preferred embodiment,an scFv that immunospecifically binds to a soluble form of B LymphocyteStimulator comprises, or alternatively consists of, the amino acidsequence of SEQ ID NOS:1570-1595. In an even more preferred embodiment,an scFv that immunospecifically binds to a soluble form of B LymphocyteStimulator comprises, or alternatively consists of, the amino acidsequence of SEQ ID NOS:1563-1569.

In another embodiment of the present invention, an scFv thatimmunospecifically binds to a membrane-bound form of B LymphocyteStimulator comprises, or alternatively consists of, the amino acidsequence of SEQ ID NOS:1881-2128 as referred to in Table 1. In apreferred embodiment, an scFv that immunospecifically binds to amembrane-bound form of B Lymphocyte Stimulator comprises, oralternatively consists of, the amino acid sequence of SEQ IDNOS:1886-1908. In an even more preferred embodiment, an scFv thatimmunospecifically binds to a membrane-bound form of B LymphocyteStimulator comprises, or alternatively consists of, the amino acidsequence of SEQ ID NOS:1881-1885.

In another embodiment of the present invention, an scFv thatimmunospecifically binds to both the soluble form and membrane-boundform of B Lymphocyte Stimulator comprises, or alternatively consists of,the amino acid sequence of SEQ ID NOS:1-1562 as referred to in Table 1.In a preferred embodiment, an scFv that immunospecifically binds to boththe soluble form and membrane-bound form of B Lymphocyte Stimulatorcomprises, or alternatively consists of, the amino acid sequence of SEQID NOS:834-872. In another preferred embodiment, an scFv thatimmunospecifically binds to both the soluble form and membrane-boundform of B Lymphocyte Stimulator comprises, or alternatively consists of,any one of the amino acids sequences of SEQ ID NOS:1-46 or 321-329.Molecules comprising, or alternatively consisting of, fragments orvariants of these scFvs, that immunospecifically bind to the solubleform of B Lymphocyte Stimulator and/or the membrane-bound form of BLymphocyte Stimulator are also encompassed by the invention, as arenucleic acid molecules encoding these scFvs, molecules, fragments and/orvariants.

In another embodiment of the present invention, scFvs thatimmunospecifically bind to the soluble form of B Lymphocyte Stimulator,comprise a polypeptide having the amino acid sequence of any one of theVH domains contained in SEQ ID NOS:1563-1880 as disclosed in Table 1and/or any one of the VL domains contained in SEQ ID NOS:1563-1880 asdisclosed in Table 1. In preferred embodiments, scFvs of the presentinvention that immunospecifically bind to the soluble form of BLymphocyte Stimulator, comprise a polypeptide having the amino acidsequence of a VH CDR and VL CDR from the same scFv referred to inTable 1. In alternative embodiments, scFvs of the present invention thatimmunospecifically bind to the soluble form of B Lymphocyte Stimulator,comprise a polypeptide having amino acid sequence of a VH CDR and VL CDRfrom different scFvs referred to in Table 1. In another embodiment,scFvs that immunospecifically bind to the soluble form of B LymphocyteStimulator, comprise a polypeptide having the amino acid sequence of anyone, two, three, or more of the VH CDRs SEQ ID NOS:1563-1880 asdisclosed in Table 1 and/or any one, two, three, or more of the VL CDRscontained in contained SEQ ID NOS:1563-1880, as disclosed in Table 1. Inpreferred embodiments, scFvs of the present invention thatimmunospecifically bind to the soluble form of B Lymphocyte Stimulator,comprise a polypeptide having the amino acid sequence of a VH domain andVL domain from the same scFv referred to in Table 1. In alternativeembodiments, scFvs of the present invention that immunospecifically bindto the soluble form of B Lymphocyte Stimulator, comprise a polypeptidehaving the of the amino acid sequence of a VH domain and VL domain fromdifferent scFvs referred to in Table 1. In a preferred embodiment, scFvsthat immunospecifically bind to the soluble form of B LymphocyteStimulator, comprise a polypeptide having the amino acid sequence of anyone of the VH CDR3s contained in SEQ ID NOS:1563-1880 as disclosed inTable 1 and/or any one of the VL CDR3s contained in SEQ ID NOS:1563-1880 as disclosed in Table 1. In preferred embodiments, scFvs ofthe present invention that immunospecifically bind to the soluble formof B Lymphocyte Stimulator, comprise a polypeptide having the amino acidsequence of a VH CDR and VL CDR from the same scFv referred to inTable 1. In alternative embodiments, scFvs of the present invention thatimmunospecifically bind to the soluble form of B Lymphocyte Stimulator,comprise a polypeptide having the amino acid sequence of a VH CDR and VLCDR from different scFvs referred to in Table 1. Molecules comprising,or alternatively consisting of, fragments or variants of these scFvs,that immunospecifically bind to B Lymphocyte Stimulator, preferably thesoluble form of B Lymphocyte Stimulator, are also encompassed by theinvention, as are nucleic acid molecules encoding these scFvs,molecules, fragments and/or variants.

In another embodiment of the present invention, scFvs thatimmunospecifically bind to the membrane-bound form of B LymphocyteStimulator comprise a polypeptide having the amino acid sequence of anyone of the VH domains contained in SEQ ID NOS:1881-2128 as disclosed inTable 1 and/or any one of the VL domains contained in SEQ ID NOS:1881-2128 as disclosed in Table 1. In preferred embodiments, scFvs ofthe present invention that immunospecifically bind to the soluble formof B Lymphocyte Stimulator, comprise a polypeptide having the amino acidsequence of a VH CDR and VL CDR from the same scFv referred to inTable 1. In alternative embodiments, scFvs of the present invention thatimmunospecifically bind to the membrane-bound form of B LymphocyteStimulator, comprise a polypeptide having the amino acid sequence of aVH domain and VL domain from different scFvs referred to in Table 1. Inanother embodiment, scFvs that immunospecifically bind to themembrane-bound form of B Lymphocyte Stimulator, comprise a polypeptidehaving the amino acid sequence of any one, two, three, or more of the VHCDRs contained in SEQ ID NOS: 1881-2128 as disclosed in Table 1 and/orany one, two, three, or more of the VL CDRs contained in SEQ ID NOS:1881-2128 as disclosed in Table 1. In preferred embodiments, scFvs ofthe present invention that immunospecifically bind to the membrane-boundform of B Lymphocyte Stimulator, comprise a polypeptide having the aminoacid sequence of a VH domain and VL domain from the same scFv referredto in Table 1. In alternative embodiments, scFvs of the presentinvention that immunospecifically bind to the membrane-bound form of BLymphocyte Stimulator, comprise a polypeptide having the amino acidsequence of a VH domain and VL domain from different scFvs referred toin Table 1. In a preferred embodiment, scFvs that immunospecificallybind to the membrane-bound form of B Lymphocyte Stimulator, comprise apolypeptide having the amino acid sequence of any one of the VH CDR3scontained in SEQ ID NOS: 1881-2128 as disclosed in Table 1 and/or anyone of the VL CDR3s contained in SEQ ID NOS: 1881-2128 as disclosed inTable 1. In preferred embodiments, scFvs of the present invention thatimmunospecifically bind to the membrane-bound form of B LymphocyteStimulator, comprise a polypeptide having the amino acid sequence of aVH domain and VL domain from the same scFv referred to in Table 1. Inalternative embodiments, scFvs of the present invention thatimmunospecifically bind to the membrane-bound form of B LymphocyteStimulator, comprise a polypeptide having the amino acid sequence of aVH CDR and VL CDR from different scFvs referred to in Table 1. Moleculescomprising, or alternatively consisting of, fragments or variants ofthese scFvs, that immunospecifically bind to B Lymphocyte Stimulator,preferably the membrane-bound form of B Lymphocyte Stimulator, are alsoencompassed by the invention, as are nucleic acid molecules encodingthese scFvs, molecules, fragments and/or variants.

In another embodiment of the present invention, scFvs thatimmunospecifically bind to the soluble form and membrane-bound form of BLymphocyte Stimulator, comprise a polypeptide having the amino acidsequence of any one of the VH domains contained in SEQ ID NOS:1-1562 asdisclosed in Table 1 and/or any one of the VL domains contained in SEQID NOS:1-1562 as disclosed in Table 1. In preferred embodiments, scFvsof the present invention that immunospecifically bind to the soluble andmembrane-bound forms of B Lymphocyte Stimulator, comprise a polypeptidehaving the amino acid sequence of a VH domain and VL domain from thesame scFv referred to in Table 1. In alternative embodiments, scFvs ofthe present invention that immunospecifically bind to the soluble formand membrane-bound form of B Lymphocyte Stimulator, comprise apolypeptide having the amino acid sequence of a VH domain and VL domainfrom different scFvs referred to in Table 1. In another embodiment,scFvs that immunospecifically bind to the soluble form andmembrane-bound form of B Lymphocyte Stimulator comprise a polypeptidehaving the amino acid sequence of any one, two, three, or more of the VHCDRs contained in SEQ ID NOS:1-1562 as disclosed in Table 1 and/or anyone, two, three, or more of the VL CDRs contained in SEQ ID NOS:1-1562as disclosed in Table 1. In preferred embodiments, scFvs of the presentinvention that immunospecifically bind to the soluble form andmembrane-bound form of B Lymphocyte Stimulator, comprise a polypeptidehaving the amino acid sequence of a VH domain and VL domain from thesame scFv referred to in Table 1. In alternative embodiments, scFvs ofthe present invention that immunospecifically bind to the soluble andmembrane-bound forms of B Lymphocyte Stimulator, comprise a polypeptidehaving the amino acid sequence of a VH domain and VL domain fromdifferent scFvs referred to in Table 1. In a preferred embodiment, scFvsthat immunospecifically bind to the soluble and membrane-bound forms ofB Lymphocyte Stimulator, comprise a polypeptide having the amino acidsequence of any one of the VH CDR3s contained in SEQ ID NOS:1-1562 asdisclosed in Table 1 and/or any one of the VL CDR3s contained in SEQ IDNOS:1-1562, as disclosed in Table 1. In preferred embodiments, scFvs ofthe present invention that immunospecifically bind to the soluble andmembrane-bound forms of B Lymphocyte Stimulator, comprise a polypeptidehaving the amino acid sequence of a VH CDR and VL CDR from the same scFvreferred to in Table 1. In alternative embodiments, scFvs of the presentinvention that immunospecifically bind to the soluble and membrane-boundforms of B Lymphocyte Stimulator, comprise a polypeptide having theamino acid sequence of a VH CDR and VL CDR from different scFvs referredto in Table 1. Molecules comprising, or alternatively consisting of,fragments or variants of these scFvs or molecules, thatimmunospecifically bind to B Lymphocyte Stimulator, preferably thesoluble and membrane-bound forms of B Lymphocyte Stimulator, are alsoencompassed by the invention, as are nucleic acid molecules encodingthese scFvs, molecules, fragments and/or variants.

The present invention provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to a polypeptide or apolypeptide fragment of B Lymphocyte Stimulator. In particular, theinvention provides antibodies corresponding to the scFvs referred to inTable 1, such scFvs may routinely be “converted” to immunoglobulinmolecules by inserting, for example, the nucleotide sequences encodingthe VH and/or VL domains of the scFv into an expression vectorcontaining the constant domain sequences and engineered to direct theexpression of the immunoglobulin molecule, as described in more detailin Example 20, infra.

In one embodiment, the invention provides antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) wherein said antibodies comprise, or alternativelyconsist of, a polypeptide having an amino acid sequence of any one ofthe VH domains contained in the sequences referred to in Table 1. Thepresent invention also provides antibodies that immunospecifically bindto a polypeptide, or polypeptide fragment of B Lymphocyte Stimulator,wherein said antibodies comprise, or alternatively consist of, apolypeptide having an amino acid sequence of any one, two, three, ormore of the VH CDRs contained in the sequences referred to in Table 1.Molecules comprising, or alternatively consisting of, these antibodies,or antibody fragments or variants thereof, that immunospecifically bindto B Lymphocyte Stimulator or a B Lymphocyte Stimulator fragment arealso encompassed by the invention, as are nucleic acid moleculesencoding these antibodies, molecules, fragments and/or variants.

In one embodiment of the present invention, antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) that immunospecifically bind B LymphocyteStimulator, comprise, or alternatively consist of, a polypeptide havingthe amino acid sequence of a VH CDR referred to in Table 1. Inparticular, the invention provides antibodies that immunospecificallybind B Lymphocyte Stimulator, comprising, or alternatively consistingof, a polypeptide having the amino acid sequence of a VH CDR1 containedin SEQ ID NOS:1-46, 321-329, 1563-1569, or 1881-1885 as disclosed inTable 1. In another embodiment, antibodies that immunospecifically bindB Lymphocyte Stimulator, comprise, or alternatively consist of, apolypeptide having the amino acid sequence of a VH CDR2 contained in SEQID NOS:1-46, 321-329, 1563-1569, or 1881-1885 as disclosed in Table 1.In a preferred embodiment, antibodies that immunospecifically bind BLymphocyte Stimulator, comprise, or alternatively consist of apolypeptide having the amino acid sequence of a VH CDR3 contained in SEQID NOS:1-46, 321-329, 1563-1569, or 1881-1885 as disclosed in Table 1.In yet another embodiment, antibodies that immunospecifically bind BLymphocyte Stimulator, comprise, or alternatively consist of, apolypeptide having the amino acid sequence of a VH CDR1 contained in SEQID NOS:834-872, 1570-1595, or 1886-1908 as disclosed in Table 1; a VHCDR2 contained in SEQ ID NOS: SEQ ID NOS: SEQ ID NOS:834-872, 1570-1595,or 1886-1908; and/or a VH CDR3 contained in SEQ ID NOS: SEQ IDNOS:834-872, 1570-1595, or 1886-1908 as disclosed in Table 1.Preferably, antibodies of the invention comprise, or alternativelyconsist of, VH CDRs that are derived from the same scFv as disclosed inTable 1. Molecules comprising, or alternatively consisting of, fragmentsor variants of these antibodies that immunospecifically bind to BLymphocyte Stimulator are also encompassed by the invention, as arenucleic acid molecules encoding these antibodies, molecules, fragmentsor variants.

The present invention provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants) that immunospecifically bind to a polypeptide, or polypeptidefragment of B Lymphocyte Stimulator. In particular, the inventionprovides antibodies wherein said antibodies comprise, or alternativelyconsist of, a VL domain having an amino acid sequence of any one of theVL domains referred to in Table 1. The present invention also providesantibodies that immunospecifically bind to a polypeptide or polypeptidefragment of B Lymphocyte Stimulator, wherein said antibodies comprise,or alternatively consist of, a VL CDR having an amino acid sequence ofany one, two, three, or more of the VL CDRs contained in the sequencesreferred to in Table 1. Molecules comprising, or alternativelyconsisting of, fragments or variants of these antibodies thatimmunospecifically bind to B Lymphocyte Stimulator are also encompassedby the invention, as are nucleic acid molecules encoding theseantibodies, molecules, fragments or variants.

In one embodiment of the present invention, antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) that immunospecifically bind B LymphocyteStimulator, comprise, or alternatively consist of, a polypeptide havingthe amino acid sequence of a VL CDR referred to in Table 1. Inparticular, the invention provides antibodies that immunospecificallybind B Lymphocyte Stimulator, comprising, or alternatively consistingof, a polypeptide having the amino acid sequence of a VL CDR1 containedin SEQ ID NOS:1-46, 321-329, 1563-1569, or 1881-1885 as disclosed inTable 1. In another embodiment, antibodies that immunospecifically bindB Lymphocyte Stimulator comprise, or alternatively consist of, apolypeptide having the amino acid sequence of a VL CDR2 contained in SEQID NOS:1-46, 321-329, 1563-1569, or 1881-1885 as disclosed in Table 1.In a preferred embodiment, antibodies comprise, or alternatively consistof, a polypeptide having the amino acid sequence of a VL CDR3 containedin SEQ ID NOS: in SEQ ID NOS:1-46, 321-329, 1563-1569, or 1881-1885disclosed in Table 1. In yet another embodiment, antibodies thatimmnospecifically bind B Lymphocyte Stimulator comprise, oralternatively consist of: a polypeptide having the amino acid sequenceof a VL CDR1 contained in SEQ ID NOS:834-872, 1570-1595, or 1886-1908 asdisclosed in Table 1; a VL CDR2 SEQ ID NOS:834-872, 1570-1595, or1886-1908 as disclosed in Table 1; and a VL CDR3 contained SEQ IDNOS:834-872, 1570-1595, or 1886-1908 as disclosed in Table 1.Preferably, antibodies of the invention comprise, or alternativelyconsist of, VL CDRs that are derived from the same scFv as disclosed inTable 1. Molecules comprising, or alternatively consisting of, fragmentsor variants of these antibodies, that immunospecifically bind to BLymphocyte Stimulator are also encompassed by the invention, as arenucleic acid molecules encoding these antibodies, molecules, fragmentsor variants.

The present invention also provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to a polypeptide or apolypeptide fragment of B Lymphocyte Stimulator, wherein said antibodiescomprise, or alternatively consist of, a VH domain of one of the scFvsreferred to in Table 1 combined with a VL domain of one of the scFvsreferred to in Table 1, or other VL domain. The present inventionfurther provides antibodies (including molecules comprise, oralternatively consist of, antibody fragments or variants thereof) thatimmunospecifically bind to a polypeptide or a polypeptide fragment of BLymphocyte Stimulator, wherein said antibodies comprise, oralternatively consist of, a VL domain of one of the scFvs referred to inTable 1 combined with a VH domain of one of the scFvs referred to inTable 1, or other VH domain. In a preferred embodiment, antibodies thatimmunospecifically bind to a polypeptide or a polypeptide fragment of BLymphocyte Stimulator, comprise, or alternatively consist of, apolypeptide having the amino acid sequence of a VH domain contained SEQID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1 and a VL domain contained in contained SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1. In a furtherpreferred embodiment, the antibodies of the invention comprise, oralternatively consist of, a VH and a VL domain from the same scFv asdisclosed in Table 1. Molecules comprising, or alternatively consistingof, fragments or variants of these antibodies, that immunospecificallybind to B Lymphocyte Stimulator are also encompassed by the invention,as are nucleic acid molecules encoding these antibodies, molecules,fragments or variants.

The present invention also provides antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants) that immunospecifically bind to a polypeptide or polypeptidefragment of B Lymphocyte Stimulator, wherein said antibodies comprise,or alternatively consist of, one, two, three, or more VH CDRs and one,two, three or more VL CDRs, as referred to in Table 1. In particular,the invention provides for antibodies that immunospecifically bind to apolypeptide or polypeptide fragment of B Lymphocyte Stimulator, whereinsaid antibodies comprise, or alternatively consist of, a VH CDR1 and aVL CDR1, a VH CDR1 and a VL CDR2, a VH CDR1 and a VL CDR3, a VH CDR2 anda VL CDR1, VH CDR2 and VL CDR2, a VH CDR2 and a VL CDR3, a VH CDR3 and aVH CDR1, a VH CDR3 and a VL CDR2, a VH CDR3 and a VL CDR3, or anycombination thereof, of the VH CDRs and VL CDRs referred to in Table 1.In a preferred embodiment, one or more of these combinations are fromthe same scFv as disclosed in Table 1. Molecules comprising, oralternatively consisting of, fragments or variants of these antibodies,that immunospecifically bind to B Lymphocyte Stimulator are alsoencompassed by the invention, as are nucleic acid molecules encodingthese antibodies, molecules, fragments or variants.

In a preferred embodiment the invention provides antibodies wherein theVH CDRX (where X=1, 2, or 3) and VL CDRY (where Y=1, 2, or 3) are fromscFvs with the same specificity (i.e., from scFvs that bind soluble BLymphocyte Stimulator, from scFvs that bind membrane-bound B LymphocyteStimulator, or from scFvs that bind both soluble and membrane-bound BLymphocyte Stimulator. Molecules comprising, or alternatively consistingof, fragments or variants of these antibodies, that immunospecificallybind to B Lymphocyte Stimulator are also encompassed by the invention,as are nucleic acid molecules encoding these antibodies, molecules,fragments or variants.

The term “antibody,” as used herein, refers to immunoglobulin moleculesand immunologically active portions of immunoglobulin molecules, i.e.,molecules that contain an antigen binding site that immunospecificallybinds an antigen. As such, the term “antibody” encompasses not onlywhole antibody molecules, but also antibody fragments, as well asvariants (including derivatives) of antibodies and antibody fragments.Antibodies of the invention include, but are not limited to, monoclonal,multispecific, human or chimeric antibodies, single chain antibodies,single chain Fvs (scFvs), Fab fragments, F(ab′)₂ fragments, Fdfragments, disulfide-linked Fvs (sdFvs), antiidiotypic (anti-Id)antibodies (including, e.g., anti-Id antibodies to antibodies of theinvention), and epitope-binding fragments of any of the above. Theimmunoglobulin molecules of the invention can be of any type (e.g., IgG,IgE, IgM, IgD, IgA and IgY), class (e.g., IgG₁, IgG₂, IgG₃, IgG₄, IgA₁and IgA₂) or subclass of immunoglobulin molecule. The antibodies of thepresent invention also include molecules comprising, or alternativelyconsisting of, a polypeptide having an amino acid sequence of a portionof an amino acid sequence contained SEQ ID NOS:1-46, 321-329, 834-872,1563-1595, or 1881-1908. Preferably, an antibody of the inventioncomprises, or alternatively consists of, a polypeptide having an aminoacid sequence of a VH domain, VH CDR, VL domain, or VL CDR of any onethose contained in the sequences referred to in Table 1. Antibodies ofthe invention also include molecules comprising, or alternativelyconsisting of, fragments or variants of the above antibodies thatimmunospecifically bind B Lymphocyte Stimulator.

Most preferably the antibodies of the present invention are wholeantibodies or antibody fragments that immunospecifically bind human BLymphocyte Stimulator. Antibody fragments of the invention thatimmunospecifically bind human B Lymphocyte Stimulator include, but arenot limited to, Fab, Fab′ and F(ab′)₂, Fd fragments, single-chain Fvs(scFv), single-chain antibodies, disulfide-linked Fvs (sdFvs), fragmentscomprising, or alternatively consisting of, either a VL or VH domain,and epitope binding fragments of any of the above.

B Lymphocyte Stimulator-binding antibody fragments, includingsingle-chain antibodies, may comprise, or alternatively consist of, thevariable region(s) alone or in combination with the entirety or aportion of the following: hinge region, CH1, CH2, and CH3 domains. In apreferred embodiment, the antibodies of the invention comprise, oralternatively consist of, a polypeptide that immunospecifically binds toB Lymphocyte Stimulator, said polypeptides comprise, or alternativelyconsist of, one, two, three, four, five, six or more CDRs referred to inTable 1, preferably a polypeptide having an amino acid sequence of a VHCDR3 and/or a VL CDR3 of contained SEQ ID NOS:1-46, 321-329, 834-872,1563-1595, or 1881-1908 as disclosed in Table 1. Most preferably,antibodies of the invention comprise, or alternatively consist of, one,two, three, four, five, six or more CDRs from the same scFv, as referredto in Table 1. The antibodies of the invention may be from any animalorigin, including birds and mammals. Preferably, the antibodies arehuman, murine (e.g., mouse and rat), donkey, sheep, rabbit, goat, guineapig, camel, horse, or chicken. Most preferably, the antibodies are humanantibodies. As used herein, “human” antibodies include antibodies havingthe amino acid sequence of a human immunoglobulin and include antibodiesisolated from human immunoglobulin libraries and xenomice or otherorganisms that have been genetically engineered to produce humanantibodies. For a detailed discussion of a few of the technologies forproducing human antibodies and human monoclonal antibodies and protocolsfor producing such antibodies, see, e.g., PCT publications WO 98/24893;WO 92/01047; WO 96/34096; WO 96/33735; European Patent No. 0 598 877;U.S. Pat. Nos. 5,413,923; 5,625,126; 5,633,425; 5,569,825; 5,661,016;5,545,806; 5,814,318; 5,885,793; 5,916,771; and 5,939,598; and Lonbergand Huszar, Int. Rev. Immunol 13:65-93 (1995), which are incorporated byreference herein in their entirety. Human antibodies or “humanized”chimeric monoclonal antibodies can be produced using techniquesdescribed herein or otherwise known in the art. For example, methods forproducing chimeric antibodies are known in the art. See, for review thefollowing references which are hereby incorporated in their entirety:Morrison, Science 229:1202 (1985); Oi et al., BioTechniques 4:214(1986); Cabilly et al., U.S. Pat. No. 4,816,567; Taniguchi et al., EP171496; Morrison et al., EP 173494; Neuberger et al., WO 8601533;Robinson et al., WO 8702671; Boulianne et al., Nature 312:643 (1984);Neuberger et al., Nature 314:268 (1985). In addition, companies such asAbgenix, Inc. (Freemont, Calif.) and Genpharm (San Jose, Calif.) can beengaged to provide human antibodies directed against a selected antigenusing technology similar to that described above.

The antibodies of the present invention may be monovalent, bivalent,trivalent or multivalent. For example, monovalent scFvs can bemultimerized either chemically or by association with another protein orsubstance. An scFv that is fused to a hexahistidine tag or a Flag tagcan be multimerized using Ni-NTA agarose (Qiagen) or using anti-Flagantibodies (Stratagene, Inc.).

The antibodies of the present invention may be monospecific, bispecific,trispecific or of greater multispecificity. Multispecific antibodies maybe specific for different epitopes of a B Lymphocyte Stimulatorpolypeptide, or fragment thereof, or may be specific for both a BLymphocyte Stimulator polypeptide, or fragment thereof, and aheterologous epitope, such as a heterologous polypeptide or solidsupport material. See, e.g., PCT publications WO 93/17715; WO 92/08802;WO 91/00360; WO 92/05793; Tutt, et al., J. Immunol. 147:60-69 (1991);U.S. Pat. Nos. 4,474,893; 4,714,681; 4,925,648; 5,573,920; 5,601,819;Kostelny et al., J. Immunol 148:1547-1553 (1992).

The antibodies of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof) maybind immunospecifically to murine B Lymphocyte Stimulator (e.g., apolypeptide having the amino acid sequence of human B LymphocyteStimulator (SEQ ID NOS:3228 and/or 3229) or B Lymphocyte Stimulatorexpressed on human monocytes; murine B Lymphocyte Stimulator (SEQ IDNOS:3230 and/or 3231) or B Lymphocyte Stimulator expressed on murinemonocytes; rat B Lymphocyte Stimulator (either the soluble forms asgiven in SEQ ID NOS:3232, 3233, 3234 and/or 3235 or in a membraneassociated form, e.g., on the surface of rat monocytes); or monkey BLymphocyte Stimulator (e.g., the monkey B Lymphocyte Stimulatorpolypeptides of SEQ ID NOS:3236 and/or 3237, the soluble form of monkeyB Lymphocyte Stimulator, or B Lymphocyte Stimulator expressed on monkeymonocytes), preferably the antibodies of the invention bindimmunospecifically to human B Lymphocyte Stimulator. Preferably, theantibodies of the invention bind immunospecifically to human and monkeyB Lymphocyte Stimulator. Also preferably, the antibodies of theinvention bind immunospecifically to human B Lymphocyte Stimulator andmurine B Lymphocyte Stimulator. More preferably, antibodies of theinvention, bind immunospecifically and with higher affinity to human BLymphocyte Stimulator than to murine B Lymphocyte Stimulator.

Antibodies of the present invention may also be described or specifiedin terms of their cross-reactivity. Antibodies that do not bind anyother analog, ortholog, or homolog of a polypeptide of the presentinvention are included. Antibodies that bind polypeptides with at least95%, at least 90%, at least 85%, at least 80%, at least 75%, at least70%, at least 65%, at least 60%, at least 55%, and at least 50% identity(as calculated using methods known in the art and described herein) to apolypeptide of the present invention are also included in the presentinvention. In a specific embodiment, antibodies of the present inventioncross react with APRIL (SEQ ID NO:3239; GenBank Accession No. AF046888;J. Exp. Med. 188(6):1185-1190; PCT International PublicationWO97/33902). In specific embodiments, antibodies of the presentinvention cross-react with murine, rat and/or rabbit homologs of humanproteins and the corresponding epitopes thereof. Antibodies that do notbind polypeptides with less than 95%, less than 90%, less than 85%, lessthan 80%, less than 75%, less than 70%, less than 65%, less than 60%,less than 55%, and less than 50% identity (as calculated using methodsknown in the art and described herein) to a polypeptide of the presentinvention are also included in the present invention. In a specificembodiment, the above-described cross-reactivity is with respect to anysingle specific antigenic or immunogenic polypeptide, or combination(s)of 2, 3, 4, 5, or more of the specific antigenic and/or immunogenicpolypeptides disclosed herein. Further included in the present inventionare antibodies which bind polypeptides encoded by polynucleotides whichhybridize to a polynucleotide of the present invention underhybridization conditions (as described herein).

In a specific embodiment, antibodies of the present invention crossreact with APRIL (SEQ ID NO:3239; GenBank Accession No. AF046888; J.Exp. Med. 188(6):1185-1190; PCT International Publication WO97/33902).In specific embodiments, antibodies that immunospecifically bind both BLymphocyte Stimulator and APRIL comprise all or a portion the BAB2001,BAB2080, BAB2015, BAB2019, BAB2087, BAB2016, BAB2034 or BAB2065 scFVs(SEQ ID NOS:3240-3247). These scFvs were isolated by panning a phagescFv library comprising VH and VL domains obtained from human bonemarrow B cells (BM library). Phage from the BM phage library were firstselected for binding to soluble B Lymphocyte Stimulator (amino acids134-285 of SEQ ID NO:3228). A second round of selection for binding tothe soluble form of APRIL (amino acids 105-250 of SEQ ID NO:3239) wasthen performed on the B Lymphocyte Stimulator binding phage selected inround one. A third round of selection for binding to the soluble form ofAPRIL (amino acids 105-250 of SEQ ID NO:3239) was then performed on thephage selected in round two. A final (fourth) round of selection forbinding to the soluble form of B Lymphocyte Stimulator (amino acids134-285 of SEQ ID NO:3228) was then performed on the phage selected inround three. Phage clones that bound B Lymphocyte Stimulator in thefourth round of selection were eluted with either 0.1M triethylamine(TEA) or with a TACI-Fc fusion protein (e.g., the extracellular domainof TACI (amino acids 31 to 159 of Genbank Accession No. AAC51790) fusedto Fc). Eluted Phage were collected and sequenced (SEQ IDNOS:3240-3247). Of 79 sequences, there were 8 unique sequences (SEQ IDNOS:3240-3247).

Isolated scFv clones (e.g., scFvs corresponding to SEQ ID NOS:3240-3247or other scFvs described in Table 1) or antibodies comprising at least aportion of said scFV clones may be screened for their ability to bindtheir ability to bind the soluble form of B Lymphocyte Stimulator andthe soluble form of APRIL by ELISA. Isolated scFv clones or antibodiescomprising at least a portion of said scFv clones may also be screenedfor their ability to inhibit binding of a soluble form of B LymphocyteStimulator or B Lymphocyte Stimulator heterotrimer to TACI, BCMA orBAFF-R (Genbank Accession Nos. AAC51790, NP_(—)00183, and NP_(—)443177,respectively). Isolated scFv clones or antibodies comprising at least aportion of said scFV clones may also be screened for their ability toinhibit B Lymphocyte Stimulator or B Lymphocyte Stimulator heterotrimermediated biological activities (e.g. stimulation of B cell proliferationand/or stimulation of immunoglobulin production).

In specific embodiments, antibodies that immunospecifically bind both BLymphocyte Stimulator and APRIL comprise all or a portion (e.g., VHCDR,VLCDR, VH domain, VL domain) of the BAB2001, BAB2015, BAB2016, BAB2019,BAB2034, BAB2065, or BAB2080 scFVs (SEQ ID NOS:3240-3247).

In preferred embodiments, the antibodies of the present invention(including molecules comprising, or alternatively consisting of,antibody fragments or variants thereof), immunospecifically bind to BLymphocyte Stimulator and do not cross-react with any other antigens. Inmore preferred embodiments, the antibodies of the inventionimmunopecifically bind to B Lymphocyte Stimulator and do not cross-reactwith TRAIL, APRIL, Endokine-alpha, TNF-alpha, TNF-beta, Fas-L or LIGHT.

The present invention also provides for a nucleic acid molecule,generally isolated, encoding an antibody of the invention (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof). In one embodiment, a nucleic acid molecule of theinvention encodes an antibody comprising, or alternatively consistingof, a VH domain having an amino acid sequence of any one of the VHdomains referred to in Table 1. In another embodiment, a nucleic acidmolecule of the present invention encodes an antibody comprising, oralternatively consisting of, a VH CDR1 having an amino acid sequence ofany one of the VH CDR1s referred to in Table 1. In another embodiment, anucleic acid molecule of the present invention encodes an antibodycomprising, or alternatively consisting of, a VH CDR2 having an aminoacid sequence of any one of the VH CDR2s referred to in Table 1. In yetanother embodiment, a nucleic acid molecule of the present inventionencodes an antibody comprising, or alternatively consisting of, a VHCDR3 having an amino acid sequence of any one of the VH CDR3s referredto in Table 1. Nucleic acid molecules encoding antibodies thatimmunospecifically bind B Lymphocyte Stimulator and comprise, oralternatively consist of, fragments or variants of the VH domains and/orVH CDRs are also encompassed by the invention.

In another embodiment, a nucleic acid molecule of the invention encodesan antibody (including molecules comprising, or alternatively consistingof, antibody fragments or variants thereof), comprising, oralternatively consisting of, a VL domain having an amino acid sequenceof any one of the VL domains referred to in Table 1. In anotherembodiment, a nucleic acid molecule of the present invention encodes anantibody comprising, or alternatively consisting of, a VL CDR1 havingamino acid sequence of any one of the VL CDR1s referred to in Table 1.In another embodiment, a nucleic acid molecule of the present inventionencodes an antibody comprising, or alternatively consisting of, a VLCDR2 having an amino acid sequence of any one of the VL CDR2s referredto in Table 1. In yet another embodiment, a nucleic acid molecule of thepresent invention encodes an antibody comprising, or alternativelyconsisting of, a VL CDR3 having an amino acid sequence of any one of theVL CDR3s referred to in Table 1. Nucleic acid encoding antibodies thatimmunospecifically bind B Lymphocyte Stimulator and comprise, oralternatively consist of, fragments or variants of the VL domains and/orVLCDR(s) are also encompassed by the invention.

In another embodiment, a nucleic acid molecule of the invention encodesan antibody (including molecules comprising, or alternatively consistingof, antibody fragments or variants thereof), comprising, oralternatively consisting of, a VH domain having an amino acid sequenceof any one of the VH domains referred to in Table 1 and a VL domainhaving an amino acid sequence of any one of the VL domains referred toin Table 1. In another embodiment, a nucleic acid molecule of theinvention encodes an antibody comprising, or alternatively consistingof, a VH CDR1, a VL CDR1, a VH CDR2, a VL CDR2, a VH CDR3, a VL CDR3, orany combination thereof having an amino acid sequence referred to inTable 1. Nucleic acid encoding antibodies that immunospecifically bind BLymphocyte Stimulator and comprise, or alternatively consist of,fragments or variants of the VL and/or domains and/or VHCDR(s) and/orVLCDR(s) are also encompassed by the invention.

The present invention also provides antibodies that comprise, oralternatively consist of, variants (including derivatives) of the VHdomains, VH CDRs, VL domains, and VL CDRs described herein, whichantibodies immunospecifically bind to B Lymphocyte Stimulator. Standardtechniques known to those of skill in the art can be used to introducemutations in the nucleotide sequence encoding a molecule of theinvention, including, for example, site-directed mutagenesis andPCR-mediated mutagenesis which result in amino acid substitutions.Preferably, the variants (including derivatives) encode less than 50amino acid substitutions, less than 40 amino acid subsitutions, lessthan 30 amino acid substitutions, less than 25 amino acid substitutions,less than 20 amino acid substitutions, less than 15 amino acidsubstitutions, less than 10 amino acid substitutions, less than 5 aminoacid substitutions, less than 4 amino acid substitutions, less than 3amino acid substitutions, or less than 2 amino acid substitutionsrelative to the reference VH domain, VHCDR1, VHCDR2, VHCDR3, VL domain,VLCDR1, VLCDR2, or VLCDR3. In specific embodiments, the variants encodesubstitutions of VHCDR3. In a preferred embodiment, the variants haveconservative amino acid substitutions at one or more predictednon-essential amino acid residues. A “conservative amino acidsubstitution” is one in which the amino acid residue is replaced with anamino acid residue having a side chain with a similar charge. Familiesof amino acid residues having side chains with similar charges have beendefined in the art. These families include amino acids with basic sidechains (e.g., lysine, arginine, histidine), acidic side chains (e.g.,aspartic acid, glutamic acid), uncharged polar side chains (e.g.,glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine),nonpolar side chains (e.g., alanine, valine, leucine, isoleucine,proline, phenylalanine, methionine, tryptophan), beta-branched sidechains (e.g., threonine, valine, isoleucine) and aromatic side chains(e.g., tyrosine, phenylalanine, tryptophan, histidine). Alternatively,mutations can be introduced randomly along all or part of the codingsequence, such as by saturation mutagenesis, and the resultant mutantscan be screened for biological activity to identify mutants that retainactivity (e.g., the ability to bind B Lymphocyte Stimulator). Followingmutagenesis, the encoded protein may routinely be expressed and thefunctional and/or biological activity of the encoded protein, (e.g.,ability to immunospecifically bind B Lymphocyte Stimulator) can bedetermined using techniques described herein or by routinely modifyingtechniques known in the art.

The antibodies of the invention include derivatives (i.e., variants)that are modified, e.g., by the covalent attachment of any type ofmolecule to the antibody such that covalent attachment does not affectthe ability of the antibody to immunospecifically bind to B LymphocyteStimulator. For example, but not by way of limitation, derivatives ofthe invention include antibodies that have been modified, e.g., byglycosylation, acetylation, pegylation, phosphorylation, amidation,derivatization by known protecting/blocking groups, proteolyticcleavage, linkage to a cellular ligand or other protein, etc. Any ofnumerous chemical modifications may be carried out by known techniques,including, but not limited to, specific chemical cleavage, acetylation,formylation, metabolic synthesis of tunicamycin, etc. Additionally, thederivative may contain one or more non-classical amino acids.

In a specific embodiment, an antibody of the invention (including amolecule comprising, or alternatively consisting of, an antibodyfragment or variant thereof), that immunospecifically binds B LymphocyteStimulator, comprises, or alternatively consists of, an amino acidsequence encoded by a nucleotide sequence that hybridizes to anucleotide sequence that is complementary to that encoding one of the VHor VL domains referred to in Table 1 under stringent conditions, e.g.,hybridization to filter-bound DNA in 6×sodium chloride/sodium citrate(SSC) at about 45° C. followed by one or more washes in 0.2×SSC/0.1% SDSat about 50-65° C., under highly stringent conditions, e.g.,hybridization to filter-bound nucleic acid in 6×SSC at about 45° C.followed by one or more washes in 0.1×SSC/0.2% SDS at about 68° C., orunder other stringent hybridization conditions which are known to thoseof skill in the art (see, for example, Ausubel, F. M. et al., eds.,1989, Current Protocols in Molecular Biology, Vol. I, Green PublishingAssociates, Inc. and John Wiley & Sons, Inc., New York at pages6.3.1-6.3.6 and 2.10.3). In another embodiment, an antibody of theinvention that immunospecifically binds to B Lymphocyte Stimulator,comprises, or alternatively consists of, an amino acid sequence encodedby a nucleotide sequence that hybridizes to a nucleotide sequence thatis complementary to that encoding one of the VH CDRs or VL CDRs referredto in Table 1 under stringent conditions, e.g., hybridization underconditions as described above, or under other stringent hybridizationconditions which are known to those of skill in the art. In anotherembodiment, an antibody of the invention that immunospecifically bindsto B Lymphocyte Stimulator, comprises, or alternatively consists of, anamino acid sequence encoded by a nucleotide sequence that hybridizes toa nucleotide sequence that is complementary to that encoding one of theVH CDR3s referred to in Table 1 under stringent conditions e.g.,hybridization under conditions as described above, or under otherstringent hybridization conditions which are known to those of skill inthe art. Nucleic acid molecules encoding these antibodies are alsoencompassed by the invention.

In another embodiment, an antibody (including a molecule comprising, oralternatively consisting of, an antibody fragment or variant thereof),that immunospecifically binds to B Lymphocyte Stimulator comprises, oralternatively consists of, a polypeptide having an amino acid sequencethat is at least 35%, at least 40%, at least 45%, at least 50%, at least55%, at least 60%, at least 65%, at least 70%, at least 75%, at least80%, at least 85%, at least 90%, at least 95%, or at least 99%identical, to any one of the VH domains referred to in Table 1. Inanother embodiment, an antibody of the invention that immunospecificallybinds to B Lymphocyte Stimulator comprises, or alternatively consistsof, a polypeptide having an amino acid sequence that is at least 35%, atleast 40%, at least 45%, at least 50%, at least 55%, at least 60%, atleast 65%, at least 70%, at least 75%, at least 80%, at least 85%, atleast 90%, at least 95%, or at least 99% identical, to any one of the VHCDRs referred to in Table 1. In another embodiment, an antibody of theinvention that immunospecifically binds to B Lymphocyte Stimulatorcomprises, or alternatively consists of, a polypeptide having an aminoacid sequence that is at least 35%, at least 40%, at least 45%, at least50%, at least 55%, at least 60%, at least 65%, at least 70%, at least75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least99% identical to any one of the VH CDR3s referred to in Table 1. Nucleicacid molecules encoding these antibodies are also encompassed by theinvention.

In another embodiment, an antibody of the invention (including amolecule comprising, or alternatively consisting of, an antibodyfragment or variant thereof), that immunospecifically binds to BLymphocyte Stimulator comprises, or alternatively consists of, apolypeptide having an amino acid sequence that is at least 35%, at least40%, at least 45%, at least 50%, at least 55%, at least 60%, at least65%, at least 70%, at least 75%, at least 80%, at least 85%, at least90%, at least 95%, or at least 99% identical, to any one of the VLdomains referred to in Table 1. In another embodiment, an antibody ofthe invention that immunospecifically binds to B Lymphocyte Stimulatorcomprises, or alternatively consists of, a polypeptide having an aminoacid sequence that is at least 35%, at least 40%, at least 45%, at least50%, at least 55%, at least 60%, at least 65%, at least 70%, at least75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least99% identical, to any one of the VL CDRs referred to in Table 1. Inanother embodiment, an antibody of the invention that immunospecificallybinds to B Lymphocyte Stimulator comprises, or alternatively consistsof, a polypeptide having an amino acid sequence that is at least 35%, atleast 40%, at least 45%, at least 50%, at least 55%, at least 60%, atleast 65%, at least 70%, at least 75%, at least 80%, at least 85%, atleast 90%, at least 95%, or at least 99% identical, to any one of the VLCDR3s referred to in Table 1. Nucleic acid molecules encoding theseantibodies are also encompassed by the invention.

Antibodies of the present invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof) mayalso be described or specified in terms of their binding affinity for toB Lymphocyte Stimulator polypeptides or fragments or variants of BLymphocyte Stimulator polypeptides (e.g., to the soluble form of BLymphocyte Stimulator and/or membrane-bound form of B LymphocyteStimulator). In specific embodiments, antibodies of the invention bind BLymphocyte Stimulator polypeptides, or fragments or variants thereof,with a dissociation constant or K_(D) of less than or equal to 5×10⁻² M,10⁻² M, 5×10⁻³ M,10⁻³ M, 5×10⁻⁴ M, 10⁻⁴ M, 5×10⁻⁵ M, or 10⁻⁵ M. Morepreferably, antibodies of the invention bind B Lymphocyte Stimulatorpolypeptides or fragments or variants thereof with a dissociationconstant or K_(D) less than or equal to 5×10⁻⁶ M, 10⁻⁶ M, 5×10⁻⁷ M, 10⁻⁷M, 5×10⁻⁸ M, or 10⁻⁸ M. Even more preferably, antibodies of theinvention bind B Lymphocyte Stimulator polypeptides or fragments orvariants thereof with a dissociation constant or K_(D) less than orequal to 5×10⁻⁹ M, 10⁻⁹ M, 5×10⁻¹⁰ M, 10⁻¹⁰ M, 5×10⁻¹¹ M, 10⁻¹¹ M,5×10⁻¹² M, 10⁻¹² M, 5×⁻¹³ M, 10⁻¹³ M, 5×10⁻¹⁴ M, 10⁻¹⁴ M, 5×10⁻¹⁵ M, or10⁻¹⁵ M. The invention encompasses antibodies that bind B LymphocyteStimulator polypeptides with a dissociation constant or K_(D) that iswithin any one of the ranges that are between each of the individualrecited values.

In specific embodiments, antibodies of the invention bind B LymphocyteStimulator polypeptides or fragments or variants thereof with an offrate (k_(off)) of less than or equal to 5×10⁻² sec⁻¹, 10⁻² sec⁻¹, 5×10⁻³sec⁻¹ or 10⁻³ sec⁻¹. More preferably, antibodies of the invention bind BLymphocyte Stimulator polypeptides or fragments or variants thereof withan off rate (k_(off)) less than or equal to 5×10⁻⁴ sec⁻¹, 10⁻⁴ sec⁻¹,5×10⁻⁵ sec⁻¹, or 10⁻⁵ sec⁻¹ 5×10⁻⁶ sec⁻¹, 10⁻⁶ sec⁻¹, 5×10⁻⁷ sec⁻¹ or10⁻⁷ sec⁻¹. The invention encompasses antibodies that bind B LymphocyteStimulator polypeptides with an off rate (k_(off)) that is within anyone of the ranges that are between each of the individual recitedvalues.

In other embodiments, antibodies of the invention bind B LymphocyteStimulator polypeptides or fragments or variants thereof with an on rate(k_(on)) of greater than or equal to 10³ M⁻¹ sec⁻¹, 5×10³ M⁻¹ sec⁻¹, 10⁴M⁻¹ sec⁻¹ or 5×10⁴ M⁻¹ sec⁻¹. More preferably, antibodies of theinvention bind B Lymphocyte Stimulator polypeptides or fragments orvariants thereof with an on rate (k_(on)) greater than or equal to 10⁵M⁻¹ sec⁻¹, 5×10⁵ M⁻¹ sec⁻¹, 10⁶ M⁻¹ sec⁻1, or 5×10⁶ M⁻¹ sec⁻¹ or 10⁷ M⁻¹sec⁻¹. The invention encompasses antibodies that bind B LymphocyteStimulator polypeptides with on rate (k_(on)) that is within any one ofthe ranges that are between each of the individual recited values.

The invention also encompasses antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that have one or more of the same biologicalcharacteristics as one or more of the antibodies described herein. By“biological characteristics” is meant, the in vitro or in vivoactivities or properties of the antibodies, such as, for example, theability to bind to B Lymphocyte Stimulator (e.g., the soluble form of BLymphocyte Stimulator, the membrane-bound form of B LymphocyteStimulator, the soluble form and membrane-bound form of B LymphocyteStimulator), and/or an antigenic and/or epitope region of B LymphocyteStimulator), the ability to substantially block B LymphocyteStimulator/B Lymphocyte Stimulator receptor (e.g., TACI—GenBankaccession number AAC51790; BCMA—GenBank accession number NP_(—)001183;and/or BAFF-R—GenBank accession number NP_(—)443177) binding, or theability to block B Lymphocyte Stimulator mediated biological activity(e.g., stimulation of B cell proliferation and immunoglobulinproduction). Optionally, the antibodies of the invention will bind tothe same epitope as at least one of the antibodies specifically referredto herein. Such epitope binding can be routinely determined using assaysknown in the art.

The present invention also provides for antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that neutralize B Lymphocyte Stimulator or a fragmentthereof, said antibodies comprising, or alternatively consisting of, aportion (i.e., a VH domain, VL domain, VH CDR1, VH CDR2, VH CDR3, VLCDR1, VL CDR2, or VL CDR3) of an scFv referred to in Table 1, morepreferably having an amino acid sequence contained in SEQ IDNOS:834-872, 1570-1595, or 1886-1908, and even more preferably having anamino acid sequence contained in SEQ ID NOS:1-46, 321-329, 1563-1569, or1881-1885 as disclosed in Table 1, or a fragment or variant thereof. Byan antibody that “neutralizes B Lymphocyte Stimulator or a fragmentthereof” is meant an antibody that diminishes or abolishes the abilityof B Lymphocyte Stimulator to bind to its receptor (e.g., TACI—GenBankaccession number AAC51790; BCMA—GenBank accession number NP 001183;and/or BAFF-R—GenBank accession number NP_(—)443177) to stimulate B cellproliferation, to stimulate immunoglobulin secretion by B cells, and/orto stimulate the B Lymphocyte Stimulator receptor signalling cascade. Inone embodiment, an antibody that neutralizes B Lymphocyte Stimulator ora fragment thereof, comprises, or alternatively consists of, apolypeptide having the amino acid sequence of a VH domain contained inSEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosedin Table 1, or a fragment or variant thereof. In another embodiment, anantibody that neutralizes B Lymphocyte Stimulator or a fragment thereof,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VL domain contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. In another embodiment, an antibody that neutralizesB Lymphocyte Stimulator or a fragment thereof, comprises, oralternatively consists of, a polypeptide having the amino acid sequenceof a VH CDR domain in SEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or1881-1908 as disclosed in Table 1, or a fragment or variant thereof. Ina preferred embodiment, an antibody that neutralizes B LymphocyteStimulator or a fragment thereof, comprises, or alternatively consistsof, a polypeptide having the amino acid sequence of a VH CDR3 containedin SEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 asdisclosed in Table 1, or a fragment or variant thereof. In anotherembodiment, an antibody that neutralizes B Lymphocyte Stimulator or afragment thereof, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VL CDR domain contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In another preferredembodiment, an antibody that neutralizes B Lymphocyte Stimulator or afragment thereof, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VL CDR3 contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. Nucleic acid moleculesencoding these antibodies are also encompassed by the invention.

The present invention also provides for antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that inhibit (i.e., diminish or abolish) B LymphocyteStimulator-mediated B cell proliferation as determined by any methodknown in the art such as, for example, the assays described in Examples21 and 22, infra, said antibodies comprising, or alternativelyconsisting of, a portion (e.g., a VH domain, VL domain, VH CDR1, VHCDR2, VH CDR3, VL CDR1, VL CDR2, or VL CDR3) of an scFv having an aminoacid sequence SEQ ID NOS:834-872, 1570-1595, 1886-1908, and even morepreferably having an amino acid sequence SEQ ID NOS:1-46, 321-329,1563-1569, 1881-1885 as disclosed in Table 1 or a fragment or variantthereof. In one embodiment, an antibody that inhibits B LymphocyteStimulator-mediated B cell proliferation, comprises, or alternativelyconsists of, a polypeptide having the amino acid sequence of a VH domaincontained in SEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908,as disclosed in Table 1, or a fragment or variant thereof. In anotherembodiment, an antibody that inhibits B Lymphocyte Stimulator-mediated Bcell proliferation, comprises, or alternatively consists of, apolypeptide having the amino acid sequence of a VL domain contained inSEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosedin Table 1, or a fragment or variant thereof. In a preferred embodiment,an antibody that inhibits B Lymphocyte Stimulator-mediated B cellproliferation, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VH CDR3 contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In another preferredembodiment, an antibody that inhibits B Lymphocyte Stimulator-mediated Bcell proliferation, comprises, or alternatively consists of, apolypeptide having the amino acid sequence of a VL CDR3 contained SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. Nucleic acid moleculesencoding these antibodies are also encompassed by the invention.

The present invention also provides for antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that inhibit (i.e., diminish or abolish) B LymphocyteStimulator-mediated stimulation of B cell survival as determined by anymethod known in the art such as, for example, the assays described inExamples 21 and 22, infra, said antibodies comprising, or alternativelyconsisting of, a portion (e.g., a VH domain, VL domain, VH CDR1, VHCDR2, VH CDR3, VL CDR1, VL CDR2, or VL CDR3) of an scFv having an aminoacid sequence SEQ ID NOS:834-872, 1570-1595, 1886-1908, and even morepreferably having an amino acid sequence SEQ ID NOS:1-46, 321-329,1563-1569, 1881-1885 as disclosed in Table 1 or a fragment or variantthereof. In one embodiment, an antibody that inhibits B LymphocyteStimulator-mediated stimulation of B cell survival, comprises, oralternatively consists of, a polypeptide having the amino acid sequenceof a VH domain contained in SEQ ID NOS:1-46, 321-329, 834-872,1563-1595, or 1881-1908, as disclosed in Table 1, or a fragment orvariant thereof. In another embodiment, an antibody that inhibits BLymphocyte Stimulator-mediated stimulation of B cell survival,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VL domain contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. In a preferred embodiment, an antibody that inhibitsB Lymphocyte Stimulator-mediated stimulation of B cell survival,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VH CDR3 contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. In another preferred embodiment, an antibody thatinhibits B Lymphocyte Stimulator-mediated stimulation of B cellsurvival, comprises, or alternatively consists of, a polypeptide havingthe amino acid sequence of a VL CDR3 contained SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. Nucleic acid molecules encoding these antibodies arealso encompassed by the invention.

The present invention also provides for antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that inhibit (i.e., diminish or abolish) B LymphocyteStimulator-mediated stimulation of B cell differentiation as determinedby any method known in the art such as, for example, the assaysdescribed in Examples 21 and 22, infra, said antibodies comprising, oralternatively consisting of, a portion (e.g., a VH domain, VL domain, VHCDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, or VL CDR3) of an scFv havingan amino acid sequence SEQ ID NOS:834-872, 1570-1595, 1886-1908, andeven more preferably having an amino acid sequence SEQ ID NOS:1-46,321-329, 1563-1569, 1881-1885 as disclosed in Table 1 or a fragment orvariant thereof. In one embodiment, an antibody that inhibits BLymphocyte Stimulator-mediated stimulation of B cell differentiation,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VH domain contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908, as disclosed in Table 1, or a fragmentor variant thereof. In another embodiment, an antibody that inhibits BLymphocyte Stimulator-mediated stimulation of B cell differentiation,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VL domain contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. In a preferred embodiment, an antibody that inhibitsB Lymphocyte Stimulator-mediated stimulation of B cell differentiation,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VH CDR3 contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. In another preferred embodiment, an antibody thatinhibits B Lymphocyte Stimulator-mediated stimulation of B celldifferentiation, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VL CDR3 contained SEQ ID NOS:1-46,321-329, 834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or afragment or variant thereof. Nucleic acid molecules encoding theseantibodies are also encompassed by the invention.

The present invention also provides for antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that inhibit (i.e., diminish or abolish) B LymphocyteStimulator-mediated stimulation of immunoglobulin production by B cellsas determined by any method known in the art such as, for example, theassays described in Examples 21 and 22, infra, said antibodiescomprising, or alternatively consisting of, a portion (e.g., a VHdomain, VL domain, VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, or VLCDR3) of an scFv having an amino acid sequence SEQ ID NOS:834-872,1570-1595, 1886-1908, and even more preferably having an amino acidsequence SEQ ID NOS:1-46, 321-329, 1563-1569, 1881-1885 as disclosed inTable 1 or a fragment or variant thereof. In one embodiment, an antibodythat inhibits B Lymphocyte Stimulator-mediated stimulation ofimmunoglobulin production by B cells, comprises, or alternativelyconsists of, a polypeptide having the amino acid sequence of a VH domaincontained in SEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908,as disclosed in Table 1, or a fragment or variant thereof. In anotherembodiment, an antibody that inhibits B Lymphocyte Stimulator-mediatedstimulation of immunoglobulin production by B cells, comprises, oralternatively consists of, a polypeptide having the amino acid sequenceof a VL domain contained in SEQ ID NOS:1-46, 321-329, 834-872,1563-1595, or 1881-1908 as disclosed in Table 1, or a fragment orvariant thereof. In a preferred embodiment, an antibody that inhibits BLymphocyte Stimulator-mediated stimulation of immunoglobulin productionby B cells, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VH CDR3 contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In another preferredembodiment, an antibody that inhibits B Lymphocyte Stimulator-mediatedstimulation of immunoglobulin production by B cells, comprises, oralternatively consists of, a polypeptide having the amino acid sequenceof a VL CDR3 contained SEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or1881-1908 as disclosed in Table 1, or a fragment or variant thereof.Nucleic acid molecules encoding these antibodies are also encompassed bythe invention.

The present invention also provides for antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that enhance the activity of B Lymphocyte Stimulatoror a fragment thereof, said antibodies comprising, or alternativelyconsisting of, a portion (i.e., a VH domain, VL domain, VH CDR1, VHCDR2, VH CDR3, VL CDR1, VL CDR2, or VL CDR3) of an scFv having an aminoacid sequence SEQ ID NOS:834-872, 1570-1595, or 1886-1908, andpreferably having an amino acid sequence of SEQ ID NOS:1-46, 321-329,1563-1569, or 1881-1885, as disclosed in Table 1, or a fragment orvariant thereof. By an antibody that “enhances the activity of BLymphocyte Stimulator or a fragment thereof” is meant an antibodyincreases the ability of B Lymphocyte Stimulator to bind to its receptor(e.g., TACI—GenBank accession number AAC51790; BCMA—GenBank accessionnumber NP_(—)001183; and/or BAFF-R—GenBank accession numberNP_(—)443177), to stimulate B cell proliferation, to stimulateimmunoglobulin secretion by B cells, and/or to stimulate the BLymphocyte Stimulator receptor signalling cascade. In one embodiment, anantibody that enhances the activity of B Lymphocyte Stimulator or afragment thereof, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VH domain contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In another embodiment, anantibody that enhances the activity of B Lymphocyte Stimulator or afragment thereof, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VL domain contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In another embodiment, anantibody that enhances the activity of B Lymphocyte Stimulator or afragment thereof, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VH CDR domain contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In a preferred embodiment, anantibody that enhances the activity of B Lymphocyte Stimulator or afragment thereof, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VH CDR3 contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In another embodiment, anantibody that enhances B Lymphocyte Stimulator or a fragment thereof,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VL CDR domain contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. In another preferred embodiment, an antibody thatenhances the activity of B Lymphocyte Stimulator or a fragment thereof,comprises, or alternatively consists of, a polypeptide having the aminoacid sequence of a VL CDR3 contained in SEQ ID NOS:1-46, 321-329,834-872, 1563-1595, or 1881-1908 as disclosed in Table 1, or a fragmentor variant thereof. Nucleic acid molecules encoding these antibodies arealso encompassed by the invention.

The present invention also provides for antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that stimulate B Lymphocyte Stimulator-mediated Bcell proliferation as determined by any method known in the art, suchas, for example, the assays described in Examples 21 and 22, infra, saidantibodies comprising, or alternatively consisting of, a portion (e.g.,a VH domain, VL domain, VH CDR1, VH CDR2, VH CDR3, VL CDR1, VL CDR2, orVL CDR3) of an scFv having an amino acid sequence of SEQ ID NOS:834-872,1570-1595, or 1886-1908, and even more preferably having an amino acidsequence of SEQ ID NOS:1-46, 321-329, 1563-1569, or 1881-1885 asdisclosed in Table 1 or a fragment or variant thereof. In oneembodiment, an antibody that stimulates B Lymphocyte Stimulator-mediatedB cell proliferation, comprises, or alternatively consists of, apolypeptide having the amino acid sequence of a VH domain contained inSEQ ID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosedin Table 1, or a fragment or variant thereof. In another embodiment, anantibody that stimulates B Lymphocyte Stimulator-mediated B cellproliferation, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VL domain contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In a preferred embodiment, anantibody that stimulates B Lymphocyte Stimulator-mediated B cellproliferation, comprises, or alternatively consists of, a polypeptidehaving the amino acid sequence of a VH CDR3 contained in SEQ IDNOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. In another preferredembodiment, an antibody that stimulates B Lymphocyte Stimulator-mediatedB cell proliferation, comprises, or alternatively consists of, apolypeptide having the amino acid sequence of a VL CDR3 contained in SEQID NOS:1-46, 321-329, 834-872, 1563-1595, or 1881-1908 as disclosed inTable 1, or a fragment or variant thereof. Nucleic acid moleculesencoding these antibodies are also encompassed by the invention.

The present invention also provides for fusion proteins comprising, oralternatively consisting of, an antibody (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that immunospecifically binds to B LymphocyteStimulator, and a heterologous polypeptide. Preferably, the heterologouspolypeptide to which the antibody is fused to is useful for B-cellfunction or is useful to target the antibody to B-cells. In analternative preferred embodiment, the heterologous polypeptide to whichthe antibody is fused to is useful for monocyte cell function or isuseful to target the antibody to a monocyte. In another embodiment, theheterologous polypeptide to which the antibody is fused is albumin(including but not limited to recombinant human serum albumin orfragments or variants thereof (see, e.g., U.S. Pat. No. 5,876,969,issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No. 5,766,883,issued Jun. 16, 1998, herein incorporated by reference in theirentirety)). In a preferred embodiment, antibodies of the presentinvention (including fragments or variants thereof) are fused with themature form of human serum albumin (i.e., amino acids 1-585 of humanserum albumin as shown in FIGS. 1 and 2 of EP Patent 0 322 094) which isherein incorporated by reference in its entirety. In another preferredembodiment, antibodies of the present invention (including fragments orvariants thereof) are fused with polypeptide fragments comprising, oralternatively consisting of, amino acid residues 1-x of human serumalbumin, where x is an integer from 1 to 585 and the albumin fragmenthas human serum albumin activity. In another preferred embodiment,antibodies of the present invention (including fragments or variantsthereof) are fused with polypeptide fragments comprising, oralternatively consisting of, amino acid residues 1-z of human serumalbumin, where z is an integer from 369 to 419, as described in U.S.Pat. No. 5,766,883 herein incorporated by reference in its entirety.Antibodies of the present invention (including fragments or variantsthereof) may be fused to either the N- or C-terminal end of theheterologous protein (e.g., immunoglobulin Fc polypeptide or human serumalbumin polypeptide).

In one embodiment, a fusion protein of the invention comprises, oralternatively consists of, a polypeptide having the amino acid sequenceof any one or more of the VH domains referred to in Table 1 or the aminoacid sequence of any one or more of the VL domains referred to in Table1 or fragments or variants thereof, and a heterologous polypeptidesequence. In another embodiment, a fusion protein of the presentinvention comprises, or alternatively consists of, a polypeptide havingthe amino acid sequence of any one, two, three, or more of the VH CDRsreferred to in Table 1, or the amino acid sequence of any one, two,three, or more of the VL CDRs referred to in Table 1, or fragments orvariants thereof, and a heterologous polypeptide sequence. In apreferred embodiment, the fusion protein comprises, or alternativelyconsists of, a polypeptide having the amino acid sequence of, a VH CDR3referred to in Table 1, or fragment or variant thereof, and aheterologous polypeptide sequence, which fusion proteinimmunospecifically binds to B Lymphocyte Stimulator. In anotherembodiment, a fusion protein comprises, or alternatively consists of apolypeptide having the amino acid sequence of at least one VH domainreferred to in Table 1 and the amino acid sequence of at least one VLdomain referred to in Table 1 or fragments or variants thereof, and aheterologous polypeptide sequence. Preferably, the VH and VL domains ofthe fusion protein correspond to the same scFv referred to in Table 1.In yet another embodiment, a fusion protein of the invention comprises,or alternatively consists of a polypeptide having the amino acidsequence of any one, two, three or more of the VH CDRs referred to inTable 1 and the amino acid sequence of any one, two, three or more ofthe VL CDRs referred to in Table 1, or fragments or variants thereof,and a heterologous polypeptide sequence. Preferably, two, three, four,five, six, or more of the VHCDR(s) or VLCDR(s) correspond to the samescFv referred to in Table 1. Nucleic acid molecules encoding thesefusion proteins are also encompassed by the invention.

The present invention also provides: antibodies (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), that immunospecifically bind to the soluble form of BLymphocyte Stimulator; antibodies that immunospecifically bind to themembrane-bound form of B Lymphocyte Stimulator; and antibodies thatimmunospecifically bind to both the soluble form and membrane-bound formof B Lymphocyte Stimulator.

In one embodiment of the present invention, antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) that immunospecifically bind to the soluble form ofB Lymphocyte Stimulator, comprise, or alternatively consist of, apolypeptide having the amino acid sequence of any one or more of the VHdomains contained in SEQ ID NOS:1563-1880 as disclosed in Table 1 and/orthe amino acid sequence of any one or more of the VL domains containedin SEQ ID NOS: 1563-1880 as disclosed in Table 1, or fragment(s) orvariant(s) (including derivative) thereof. Preferably, the VH and VLdomains of the antibody correspond to the same scFv as disclosed inTable 1. In another embodiment, antibodies that immunospecifically bindto the soluble form of B Lymphocyte Stimulator are provided thatcomprise, or alternatively consist of, a polypeptide having the aminoacid sequence of any one, two, three, or more of the VH CDRs containedSEQ ID NOS: 1563-1880 as disclosed in Table 1 and/or the amino acidsequence of any one, two, three, or more of the VL CDRs contained in SEQID NOS: 1563-1880 as disclosed in Table 1, or fragment(s) or variant(s)thereof. Preferably, two, three, four, five, six or more of the VH andVL CDRs of the antibody correspond to the same scFv as disclosed inTable 1. In a preferred embodiment, antibodies that immunospecificallybind to the soluble form of B Lymphocyte Stimulator are provided thatcomprise, or alternatively consist of, a polypeptide having the aminoacid sequence of any one or more of the VH CDR3s contained in SEQ IDNOS: 1563-1880 as disclosed in Table 1 and/or the amino acid sequence ofany one or more of the VL CDR3s contained in SEQ ID NOS: 1563-1880 asdisclosed in Table 1, or fragment(s) or variant(s) thereof. Preferably,the VHCDR3 and VLCDR3 of the antibody correspond to the same scFv, asdisclosed in Table 1. Nucleic acid molecules encoding these antibodiesare also encompassed by the invention.

In another embodiment of the present invention, antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) that immunospecifically bind to the membrane-boundform of B Lymphocyte Stimulator are provided that comprise, oralternatively consist of, a polypeptide having the amino acid sequenceof any one or more of the VH domains contained in SEQ ID NOS: 1881-2128as disclosed in Table 1 and/or the amino acid sequence of any one ormore of the VL domains contained in SEQ ID NOS: 1881-2128 as disclosedin Table 1, or a fragment or variant thereof. Preferably, the VH and VLdomains of the antibody correspond to the same scFv as disclosed inTable 1. In another embodiment, antibodies that immunospecifically bindto the membrane-bound form of B Lymphocyte Stimulator are provided thatcomprise, or alternatively consist of, a polypeptide having the aminoacid sequence of any one, two, three, or more of the VH CDRs containedin SEQ ID NOS: 1881-2128 as disclosed in Table 1 and/or the amino acidsequence of any one, two, three, or more of the VL CDRs contained in SEQID NOS: 1881-2128 as disclosed in Table 1, or fragment(s) or variant(s)thereof. Preferably, two, three, four, five, six or more of the VH andVL CDRs of the antibody correspond to the same scFv as disclosed inTable 1. In a preferred embodiment, antibodies that immunospecificallybind to the membrane-bound form of B Lymphocyte Stimulator are providedthat comprise, or alternatively consist of, a polypeptide having theamino acid sequence of any one or more of the VH CDR3s contained in SEQID NOS: 1881-2128 as disclosed in Table 1 and/or the amino acid sequenceof any one or more of the VL CDR3s contained in SEQ ID NOS: 1881-2128 asdisclosed in Table 1, or fragment(s) or variant(s) thereof. Preferably,the VHCDR3 and VLCDR3 of the antibody correspond to the same scFv, asdisclosed in Table 1. Nucleic acid molecules encoding these antibodiesare also encompassed by the invention.

In another embodiment of the present invention, antibodies (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) that immunospecifically bind to the soluble formand membrane-bound form of B Lymphocyte Stimulator, are provided thatcomprise, or alternatively consist of, a polypeptide having the aminoacid sequence of any one or more of the VH domains contained in SEQ IDNOS: 1-1562 as disclosed in Table 1 and/or the amino acid sequence ofany one or more of the VL domains contained in SEQ ID NOS: 1-1562 asdisclosed in Table 1, or a fragment or variant thereof. Preferably, theVH and VL domains of the antibody correspond to the same scFv asdisclosed in Table 1. In another embodiment, antibodies thatimmunospecifically bind to the soluble form and membrane-bound form of BLymphocyte Stimulator are provided that comprise, or alternativelyconsist of, a polypeptide having the amino acid sequence of any one,two, three, or more of the VH CDRs contained in SEQ ID NOS: 1-1562 asdisclosed in Table 1 and/or the amino acid sequence of any one, two,three, or more of the VL CDRs contained in SEQ ID NOS: 1-1562 asdisclosed in Table 1, or fragment(s) or variant(s) thereof. Preferably,two, three, four, five, six or more of the VH and VL CDRs of theantibody correspond to the same scFv as disclosed in Table 1. In apreferred embodiment, antibodies that immunospecifically bind to thesoluble form and membrane-bound form of B Lymphocyte Stimulator areprovided that comprise, or alternatively consist of, a polypeptidehaving the amino acid sequence of any one or more of the VH CDR3scontained in SEQ ID NOS: 1-1562, disclosed in Table 1 and/or the aminoacid sequence of any one or more of the VL CDR3s contained in SEQ IDNOS: 1-1562, disclosed in Table 1, or fragment(s) or variant(s) thereof.Preferably, the VHCDR3 and VLCDR3 of the antibody correspond to the samescFv, as disclosed in Table 1.

The present invention also provides for mixtures of antibodies(including scFvs and other molecules comprising, or alternativelyconsisting of, antibody fragments or variants thereof) thatimmunospecifically bind to B Lymphocyte Stimulator, wherein the mixturehas at least one, two, three, four, five or more different antibodies ofthe invention. In particular, the invention provides for mixtures ofdifferent antibodies that immunospecifically bind to the soluble form ofB Lymphocyte Stimulator, the membrane-bound form of B LymphocyteStimulator, and/or both the membrane-bound form and soluble form of BLymphocyte Stimulator. In specific embodiments, the invention providesmixtures of at least 2, preferably at least 4, at least 6, at least 8,at least 10, at least 12, at least 15, at least 20, or at least 25different antibodies that immunospecifically bind to B LymphocyteStimulator, wherein at least 1, at least 2, at least 4, at least 6, orat least 10, antibodies of the mixture is an antibody of the invention.In a specific embodiment, each antibody of the mixture is an antibody ofthe invention.

The present invention also provides for panels of antibodies (includingscFvs and other molecules comprising, or alternatively consisting of,antibody fragments or variants thereof) that immunospecifically bind toB Lymphocyte Stimulator, wherein the panel has at least one, two, three,four, five or more different antibodies of the invention. In particular,the invention provides for panels of different antibodies thatimmunospecifically bind to the soluble form of B Lymphocyte Stimulator,the membrane-bound form of B Lymphocyte Stimulator, and/or both themembrane-bound form and soluble form of B Lymphocyte Stimulator. Inspecific embodiments, the invention provides for panels of antibodiesthat have different affinities for B Lymphocyte Stimulator, differentspecificities for B Lymphocyte Stimulator, or different dissociationrates. The invention provides panels of at least 10, preferably at least25, at least 50, at least 75, at least 100, at least 125, at least 150,at least 175, at least 200, at least 250, at least 300, at least 350, atleast 400, at least 450, at least 500, at least 550, at least 600, atleast 650, at least 700, at least 750, at least 800, at least 850, atleast 900, at least 950, or at least 1000, antibodies. Panels ofantibodies can be used, for example, in 96 well plates for assays suchas ELISAs.

The present invention further provides for compositions comprising, oneor more antibodies (including scFvs and other molecules comprising, oralternatively consisting of antibody fragments or variants of theinvention). In one embodiment, a composition of the present inventioncomprises, one, two, three, four, five, or more antibodies that compriseor alternatively consist of, a polypeptide having an amino acid sequenceof any one or more of the VH domains contained in SEQ ID NOS:1563-1880as disclosed in Table 1, or a variant thereof. In another embodiment, acomposition of the present invention comprises, one, two, three, four,five, or more antibodies that comprise, or alternatively consist of, apolypeptide having an amino acid sequence of any one or more of the VHCDR1s contained in SEQ ID NOS:1563-1880 as disclosed in Table 1, or avariant thereof. In another embodiment, a composition of the presentinvention comprises, one, two, three, four, five or more antibodies thatcomprise, or alternatively consist of, a polypeptide having an aminoacid sequence of any one or more of the VH CDR2s contained in SEQ IDNOS:1563-1880 as disclosed in Table 1, or a variant thereof. In apreferred embodiment, a composition of the present invention comprises,one, two, three, four, five, or more antibodies that comprise, oralternatively consist of, a polypeptide having an amino acid sequence ofany one or more of the VH CDR3s contained in SEQ ID NOS:1563-1880, asdisclosed in Table 1 or a variant thereof.

The present invention further provides for compositions comprising, oneor more antibodies (including scFvs and other molecules comprising, oralternatively consisting of antibody fragments or variants of theinvention). In one embodiment, a composition of the present inventioncomprises, one, two, three, four, five, or more antibodies that compriseor alternatively consist of, a polypeptide having an amino acid sequenceof any one or more of the VH domains contained in SEQ ID NOS:1881-2128as disclosed in Table 1, or a variant thereof. In another embodiment, acomposition of the present invention comprises, one, two, three, four,five, or more antibodies that comprise, or alternatively consist of, apolypeptide having an amino acid sequence of any one or more of the VHCDR1s contained in SEQ ID NOS:1881-2128 as disclosed in Table 1, or avariant thereof. In another embodiment, a composition of the presentinvention comprises, one, two, three, four, five or more antibodies thatcomprise, or alternatively consist of, a polypeptide having an aminoacid sequence of any one or more of the VH CDR2s contained in SEQ IDNOS:1881-2128 as disclosed in Table 1, or a variant thereof. In apreferred embodiment, a composition of the present invention comprises,one, two, three, four, five, or more antibodies that comprise, oralternatively consist of, a polypeptide having an amino acid sequence ofany one or more of the VH CDR3s contained in SEQ ID NOS:1881-2128 asdisclosed in Table 1 or a variant thereof.

The present invention further provides for compositions comprising, oneor more antibodies (including scFvs, or molecules comprising, oralternatively consisting of antibody fragments or variants of theinvention). In one embodiment, a composition of the present inventioncomprises, one, two, three, four, five, or more antibodies that compriseor alternatively consist of, a polypeptide having an amino acid sequenceof any one or more of the VH domains contained in SEQ ID NOS:1-1562 asdisclosed in Table 1, or a variant thereof. In another embodiment, acomposition of the present invention comprises, one, two, three, four,five, or more antibodies that comprise, or alternatively consist of, apolypeptide having an amino acid sequence of any one or more of the VHCDR1s contained in SEQ ID NOS:1-1562 as disclosed in Table 1, or avariant thereof. In another embodiment, a composition of the presentinvention comprises, one, two, three, four, five or more antibodies thatcomprise, or alternatively consist of, a polypeptide having an aminoacid sequence of any one or more of the VH CDR2s contained in SEQ IDNOS:1-1562 as disclosed in Table 1, or a variant thereof. In a preferredembodiment, a composition of the present invention comprises, one, two,three, four, five, or more antibodies that comprise, or alternativelyconsist of, a polypeptide having an amino acid sequence of any one ormore of the VH CDR3s contained in SEQ ID NOS:1-1562 as disclosed inTable 1 or a variant thereof.

Other embodiments of the present invention providing for compositionscomprising, one or more antibodies (including scFvs and other moleculescomprising, or alternatively consisting of antibody fragments orvariants of the invention) are listed below. In another embodiment, acomposition of the present invention comprises, one, two, three, four,five, or more antibodies that comprise, or alternative consist of, apolypeptide having an amino acid sequence of any one or more of the VLdomains contained in SEQ ID NOS:1563-1880 as disclosed in Table 1, or avariant thereof. In another embodiment, a composition of the presentinvention comprises, one, two, three, four, five, or more antibodiesthat comprise, or alternatively consist of, a polypeptide having anamino acid sequence of any one or more of the VL CDR1s contained in SEQID NOS:1563-1880 as disclosed in Table 1, or a variant thereof. Inanother embodiment, a composition of the present invention comprises,one, two, three, four, five, or more antibodies that comprise, oralternatively consist of, a polypeptide having an amino acid sequence ofany one or more of the VL CDR2s contained SEQ ID NOS:1563-1880 asdisclosed in Table 1, or a variant thereof. In a preferred embodiment, acomposition of the present invention comprises, one, two, three, four,five, or more antibodies that comprise, or alternatively consist of, apolypeptide having an amino acid sequence of any one or more of the VLCDR3s contained in SEQ ID NOS:1563-1880 as disclosed in Table 1, or avariant thereof.

Other embodiments of the present invention providing for compositionscomprising, one or more antibodies (including scFvs and other moleculescomprising, or alternatively consisting of antibody fragments orvariants of the invention) are listed below. In another embodiment, acomposition of the present invention comprises, one, two, three, four,five, or more antibodies that comprise, or alternatively consist of, apolypeptide having an amino acid sequence of any one or more of the VLdomains contained in SEQ ID NOS:1881-2128 as disclosed in Table 1, or avariant thereof. In another embodiment, a composition of the presentinvention comprises, one, two, three, four, five, or more antibodiesthat comprise, or alternatively consist of, a polypeptide having anamino acid sequence of any one or more of the VL CDR1s contained in SEQID NOS:1881-2128 as disclosed in Table 1, or a variant thereof. Inanother embodiment, a composition of the present invention comprises,one, two, three, four, five, or more antibodies that comprise, oralternatively consist of, a polypeptide having an amino acid sequence ofany one or more of the VL CDR2s SEQ ID NOS:1881-2128 as disclosed inTable 1, or a variant thereof. In a preferred embodiment, a compositionof the present invention comprises, one, two, three, four, five, or moreantibodies that comprise, or alternatively consist of, a polypeptidehaving an amino acid sequence of any one or more of the VL CDR3scontained in SEQ ID NOS:1881-2128 as disclosed in Table 1, or a variantthereof.

Other embodiments of the present invention providing for compositionscomprising, one or more antibodies (including scFvs and other moleculescomprising, or alternatively consisting of antibody fragments orvariants of the invention) are listed below. In another embodiment, acomposition of the present invention comprises, one, two, three, four,five, or more antibodies that comprise, or alternatively consist of, apolypeptide having an amino acid sequence of any one or more of the VLdomains contained in SEQ ID NOS:1-1562 as disclosed in Table 1, or avariant thereof. In another embodiment, a composition of the presentinvention comprises, one, two, three, four, five, or more antibodiesthat comprise, or alternatively consist of, a polypeptide having anamino acid sequence of any one or more of the VL CDR1s contained in SEQID NOS:1-1562 as disclosed in Table 1, or a variant thereof. In anotherembodiment, a composition of the present invention comprises, one, two,three, four, five, or more antibodies that comprise, or alternativelyconsist of, a polypeptide having an amino acid sequence of any one ormore of the VL CDR2s SEQ ID NOS:1-1562 as disclosed in Table 1, or avariant thereof. In a preferred embodiment, a composition of the presentinvention comprises, one, two, three, four, five, or more antibodiesthat comprise, or alternatively consist of, a polypeptide having anamino acid sequence of any one or more of the VL CDR3s contained in SEQID NOS:1-1562 as disclosed in Table 1, or a variant thereof.

In a preferred embodiment, a composition of the present inventioncomprises, one, two, three, four, five, or more antibodies thatcomprise, or alternatively consist of, a polypeptide having an aminoacid sequence of any one or more of the VH domains in disclosed in Table1, or a variant thereof, and an amino acid sequence of any one or moreof the VL domains disclosed in Table 1, or a variant thereof wherein theVH and VL domains are from scFvs with the same specificity (i.e., fromscFvs that bind soluble B Lymphocyte Stimulator (SEQ ID NOS:1563-1880),from scFvs that bind membrane-bound B Lymphocyte Stimulator (SEQ ID1881-2128), or from scFvs that bind both soluble and membrane-bound BLymphocyte Stimulator (SEQ ID NOS:1-1562). In a preferred embodiment theinvention provides antibodies wherein the VH CDRX (where X=1,2, or 3)and VL CDRY (where Y=1,2, or 3) are from scFvs with the same specificity(i.e., from scFvs that bind soluble B Lymphocyte Stimulator (SEQ IDNOS:1563-1880), from scFvs that bind membrane-bound B LymphocyteStimulator (SEQ ID NOS:1881-2128), or from scFvs that bind both solubleand membrane-bound B Lymphocyte Stimulator (SEQ ID NOS:1-1562). In yetanother embodiment, a composition of the present invention comprises oneor more fusion proteins.

As discussed in more detail below, a composition of the invention may beused either alone or in combination with other compositions. Theantibodies (including scFvs and other molecules comprising, oralternatively consisting of antibody fragments or variants of thepresent invention) may further be recombinantly fused to a heterologouspolypeptide at the N- or C-terminus or chemically conjugated (includingcovalently and non-covalently conjugations) to polypeptides or othercompositions. For example, antibodies of the present invention may berecombinantly fused or conjugated to molecules useful as labels indetection assays and effector molecules such as heterologouspolypeptides, drugs, radionuclides, or toxins. See, e.g., PCTpublications WO 92/08495; WO 91/14438; WO 89/12624; U.S. Pat. No.5,314,995; and EP 396,387.

Antibodies of the present invention (including scFvs and other moleculescomprising, or alternatively consisting of antibody fragments orvariants of the present invention) may be used, for example, but notlimited to, to purify and detect B Lymphocyte Stimulator, and to targetthe polypeptides of the present invention to cells expressingmembrane-bound B Lymphocyte Stimulator or B Lymphocyte Stimulatorreceptor, including both in vitro and in vivo diagnostic and therapeuticmethods. For example, the antibodies have use in immunoassays forqualitatively and quantitatively measuring levels of B LymphocyteStimulator in biological samples. See, e.g., Harlow et al., Antibodies:A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988)(incorporated by reference herein in its entirety).

Methods Producing Antibodies

The antibodies of the invention (including scFvs and other moleculescomprising, or alternatively consisting of antibody fragments orvariants of the invention) can be produced by any method known in theart for the synthesis of antibodies, in particular, by chemicalsynthesis or preferably, by recombinant expression techniques.

The single chain Fvs disclosed in Table 1 were generated using phagedisplay methods known in the art. Furthermore, other scFvs thatimmunospecifically bind B Lymphocyte Stimulator may be generated usingphage display methods known in the art. In phage display methods,functional antibody domains are displayed on the surface of phageparticles which carry the polynucleotide sequences encoding them. Inparticular, DNA sequences encoding VH and VL domains are amplified fromanimal cDNA libraries (e.g., human or murine cDNA libraries of lymphoidtissues) or synthetic cDNA libraries. The DNA encoding the VH and VLdomains are joined together by an scFv linker by PCR and cloned into aphagemid vector (e.g., p CANTAB 6 or pComb 3 HSS). The vector iselectroporated in E. coli and the E. coli is infected with helper phage.Phage used in these methods are typically filamentous phage including fdand M13 and the VH and VL domains are usually recombinantly fused toeither the phage gene III or gene VIII. Phage expressing an antigenbinding domain that binds to an antigen of interest (i.e., B LymphocyteStimulator or a fragment thereof) can be selected or identified withantigen, e.g., using labeled antigen or antigen bound or captured to asolid surface or bead. Examples of phage display methods that can beused to make the antibodies of the present invention include, but arenot limited to, those disclosed in Brinkman et al., J. Immunol. Methods182:41-50 (1995); Ames et al., J. Immunol. Methods 184:177-186 (1995);Kettleborough et al., Eur. J. Immunol. 24:952-958 (1994); Persic et al.,Gene 187 9-18 (1997); Burton et al., Advances in Immunology57:191-280(1994); PCT application No. PCT/GB91/O1 134; PCT publicationsWO 90/02809; WO 91/10737; WO 92/01047; WO 92/18619; WO 93/1 1236; WO95/15982; WO 95/20401; WO97/13844; and U.S. Pat. Nos. 5,698,426;5,223,409; 5,403,484; 5,580,717; 5,427,908; 5,750,753; 5,821,047;5,571,698; 5,427,908; 5,516,637; 5,780,225; 5,658,727; 5,733,743 and5,969,108; each of which is incorporated herein by reference in itsentirety.

ScFvs that immunospecifcally bind to both B Lymphocyte Stimulator andAPRIL polypeptides (preferably to the mature soluble forms of each) maybe obtained, for example, by sequential rounds of selection for bindingto one or the other of B Lymphocyte Stimulator and APRIL polypeptides.Thus in one embodiment, the present invention provides for a method ofselecting phage that express scFvs that immunospecifically bind both BLymphocyte Stimulator polypeptides and APRIL polypeptides, comprising atleast one round of phage selection for binding to B LymphocyteStimulator polypeptide and at least one round of phage selection forbinding to APRIL polypeptide. Selection for B Lymphocyte Stimulatorbinding may either precede or follow selection for APRIL binding. Morethan one round of selection for binding to either B LymphocyteStimulator or APRIL may be conducted.

ScFvs that immunospecifcally bind to a heterotrimer comprising at leastone B Lymphocyte Stimulator polypeptide and at least one APRILpolypeptide may be, for example, obtained by sequential rounds ofselection for binding to one or the other of B Lymphocyte Stimulator andAPRIL polypeptide. Thus in one embodiment, the present inventionprovides for a method of selecting phage that express scFvs thatimmunospecifically bind a heterotrimer comprising at least one BLymphocyte Stimulator polypeptide and at least one APRIL polypeptide,comprising at least one round of phage selection for binding to BLymphocyte Stimulator polypeptide and at least one round of phageselection for binding to APRIL polypeptide. Selection for B LymphocyteStimulator binding may either precede or follow selection for APRILbinding. More than one round of selection for binding to either BLymphocyte Stimulator or APRIL may be conducted.

Alternatively, scFvs that immunospecifcally bind to a heterotrimercomprising at least one B Lymphocyte Stimulator polypeptide and at leastone APRIL polypeptide may be obtained, for example, by selecting scFVsthat bind to a B Lymphocyte Stimulator heterotrimer. A B LymphocyteStimulator heterotrimer may contain, for example, one B LymphocyteStimulator polypeptide and two APRIL polypeptides (2B LymphocyteStimulator:1APRIL heterotrimer). Other B Lymphocyte Stimulatorheterotrimers may contain, for example, one B Lymphocyte Stimulatorpolypeptide and two APRIL polypeptides (1B Lymphocyte Stimulator:2APRILheterotrimer). Preferably, the heterotrimers comprising B LymphocyteStimulator and APRIL polypeptides contain the mature forms of both the BLymphocyte Stimulator and APRIL polypeptides. ScFvs may be selected inone or more rounds of selection for binding only to the 2B LymphocyteStimulator:1APRIL heterotrimer, or only to the 1B LymphocyteStimulator:2APRIL heterotrimer. Alternatively, scFVs thatimmunospecifcally bind heterotrimers comprising B Lymphocyte Stimulatorand APRIL polypeptides may be obtained through sequential rounds ofscreening on individual forms of the B Lymphocyte Stimulator/APRILheterotrimer in any order and/or on both forms of the heterotrimersimultaneously, or any combination thereof.

As described in the above references, after phage selection, theantibody coding regions from the phage can be isolated and used togenerate whole antibodies, including human antibodies, or any otherdesired antigen binding fragment, and expressed in any desired host,including mammalian cells, insect cells, plant cells, yeast, andbacteria, e.g., as described below. Techniques to recombinantly produceFab, Fab′ and F(ab′)2 fragments can also be employed using methods knownin the art such as those disclosed in PCT publication WO 92/22324;Mullinax et al., BioTechniques 12(6):864-869 (1992); Sawai et al., AJRI34:26-34 (1995); and Better et al., Science 240:1041-1043 (1988) (saidreferences incorporated by reference in their entireties).

The characteristics of an antibody, such as its on-rate, off-rate and/oroverall affinity may be altered using in vitro mutation and selectiontechniques. This process is well known in the art and is commonlyreferred to as in vitro affinity maturation of antibodies. Starting witha given antibody that binds a particular antigen with a certainaffinity, one of skill in the art can engineer variants of that antibodyand test the antibody variants for altered (usually improved) antigenbinding characteristics. The amino acid sequence of the VH and VLregions of such antibody variants may be substantially from that of thestarting or original antibody; for example, the an antibody variants maycomprise the original VH or VL paired with a different (from theoriginal) VL or VH region, respectively. Alternatively, the amino acidsequence of the VH and VL region of such antibody variants may be quitesimilar to that of the starting or original antibody; for example, anantibody variant may have only a few amino acid changes in the VH and/orVL region. It is common for one to engineer the mutations in CDRregions, and particularly in theVHCDR3 region. Examples of both types ofin vitro antibody affinity maturation are described for example,Thompson et al., (1996) The Journal of Molecular Biology 256:77-88.Moreover, a review of phage display antibody technology may be found inVaughan et al., (1998) Nature Biotechnology 16:535-539; both of thesearticles are herein incorporated by reference in their entireties. ThescFvs of SEQ ID NOS:10-37 are CDR3 mutants derived from the scFv of SEQID NO:9 and scFvs of SEQ ID NOS:291-327 are CDR3 mutants derived fromthe scFv of SEQ ID NO:2.

To generate whole antibodies, PCR primers including VH or VL nucleotidesequences, a restriction site, and a flanking sequence to protect therestriction site can be used to amplify the VH or VL sequences in scFvclones. Utilizing cloning techniques known to those of skill in the art,the PCR amplified VH domains can be cloned into vectors expressing a VHconstant region, e.g., the human gamma 4 constant region, and the PCRamplified VL domains can be cloned into vectors expressing a VL constantregion, e.g., human kappa or lambda constant regions. Preferably, thevectors for expressing the VH or VL domains comprise a promoter suitableto direct expression of the heavy and light chains in the chosenexpression system, a secretion signal, a cloning site for theimmunoglobulin variable domain, immunoglobulin constant domains, and aselection marker such as neomycin. The VH and VL domains may also becloned into one vector expressing the necessary constant regions. Theheavy chain conversion vectors and light chain conversion vectors arethen co-transfected into cell lines to generate stable or transient celllines that express full-length antibodies, e.g., IgG, using techniquesknown to those of skill in the art.

Cell lines that express antibodies that comprise the VH and VL domainsof scFvs of the invention have been deposited with the American TypeCulture Collection (”ATCC™”) on the dates listed in Table 2 and giventhe ATCC™ Deposit Numbers identified in Table 2. The ATCC™ is located at10801 University Boulevard, Manassas, Va. 20110-2209, USA. The ATCC™deposit was made pursuant to the terms of the Budapest Treaty on theinternational recognition of the deposit of microorganisms for purposesof patent procedure.

TABLE 2 ATCC ™ ATCC ™ Corresponding SEQ ID Deposit Deposit Cell LinescFv NO: Number Date NSO-B11-15 I050B11-15 24 PTA-3238 Mar. 27, 2001NSO-anti-B I006D08 2 PTA-3239 Mar. 27, 2001 Lymphocyte Stimulator-6D08-18 NSO-anti-B I116A01 327 PTA-3240 Mar. 27, 2001 LymphocyteStimulator- 116A01-60 IO26C04K I026C04-K 1563 PTA-3241 Mar. 27, 2001IO50A12 I050A12 12 PTA-3242 Mar. 27, 2001 IO50-B11 I050B11 9 PTA-3243Mar. 27, 2001

Accordingly, in one embodiment, the invention provides antibodies thatcomprise the VH and VL domains of scFvs of the invention.

In a preferred embodiment, an antibody of the invention is the antibodyexpressed by cell line NSO-B11-15.

In a preferred embodiment, an antibody of the invention is the antibodyexpressed by cell line NSO-anti-B Lymphocyte Stimulator-6D08-18.

In a preferred embodiment, an antibody of the invention is the antibodyexpressed by cell line NSO-anti-B Lymphocyte Stimulator-116A01-60.

In a preferred embodiment, an antibody of the invention is the antibodyexpressed by cell line IO26C04K.

In a preferred embodiment, an antibody of the invention is the antibodyexpressed by cell line IO50A12.

In a preferred embodiment, an antibody of the invention is the antibodyexpressed by cell line NSO-B11.

In other preferred embodiments, the invention provides antibodies thatcompetitively inhibit binding of an antibody comprising a fragment(e.g., VH domain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, orVLCDR3) or variant of an scFv referred to in Table 1 to a B LymphocyteStimulator polypeptide. In preferred embodiments, the invention providesantibodies that which reduce the binding of an antibody comprising afragment (e.g., VH domain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1,VLCDR2, or VLCDR3) or variant of an scFv referred to in Table 1 to a BLymphocyte Stimulator polypeptide by between 1% and 10% in a competitiveinhibition assay. In preferred embodiments, the invention providesantibodies that which reduce the binding of an antibody comprising afragment (e.g., VH domain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1,VLCDR2, or VLCDR3) or variant of an scFv referred to in Table 1 to a BLymphocyte Stimulator polypeptide by between 1% and 10% in a competitiveinhibition assay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 10% and up to 20% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 20% and up to 30% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 30% and up to 40% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 40% and up to 50% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 50% and up to 60% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 60% and up to 70% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 70% and up to 80% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 80% and up to 90% in a competitive inhibitionassay.

In preferred embodiments, the invention provides antibodies that whichreduce the binding of an antibody comprising a fragment (e.g., VHdomain, VL domain, VHCDR1, VHCDR2, VHCDR3, VLCDR1, VLCDR2, or VLCDR3) orvariant of an scFv referred to in Table 1 to a B Lymphocyte Stimulatorpolypeptide by at least 90% and up to 100% in a competitive inhibitionassay.

In other preferred embodiments, the invention provides antibodies thatcompetitively inhibit binding of the antibody produced by the cell linehaving ATCC™ deposit number PTA-3238 to a B Lymphocyte Stimulatorpolypeptide.

In other preferred embodiments, the invention provides antibodies thatcompetitively inhibit binding of the antibody produced by the cell linehaving ATCC™ deposit number PTA-3239 to a B Lymphocyte Stimulatorpolypeptide.

In other preferred embodiments, the invention provides antibodies thatcompetitively inhibit binding of the antibody produced by the cell linehaving ATCC™ deposit number PTA-3240 to a B Lymphocyte Stimulatorpolypeptide.

In other preferred embodiments, the invention provides antibodies thatcompetitively inhibit binding of the antibody produced by the cell linehaving ATCC™ deposit number PTA-3241 to a B Lymphocyte Stimulatorpolypeptide.

In other preferred embodiments, the invention provides antibodies thatcompetitively inhibit binding of the antibody produced by the cell linehaving ATCC™ deposit number PTA-3242 to a B Lymphocyte Stimulatorpolypeptide.

In other preferred embodiments, the invention provides antibodies thatcompetitively inhibit binding of the antibody produced by the cell linehaving ATCC™ deposit number PTA-3243 to a B Lymphocyte Stimulatorpolypeptide.

For some uses, including in vivo use of antibodies in humans and invitro detection assays, it may be preferable to use human or chimericantibodies. Completely human antibodies are particularly desirable fortherapeutic treatment of human patients. See also, U.S. Pat. Nos.4,444,887 and 4,716,111; and PCT publications WO 98/46645, WO 98/50433,WO 98/24893, WO98/16654, WO 96/34096, WO 96/33735, and WO 91/10741; eachof which is incorporated herein by reference in its entirety. In aspecific embodiment, antibodies of the present invention comprise one ormore VH and VL domains corresponding to the human scFvs of the inventionand framework regions from another immunoglobulin molecule, preferably ahuman immunoglobulin molecule. In a specific embodiment, antibodies ofthe present invention comprise one or more CDRs corresponding to thehuman scFvs of the invention and framework regions from anotherimmunoglobulin molecule, preferably a human immunoglobulin molecule. Inother embodiments, an antibody of the present invention comprises one,two, three, four, five, six or more VL CDRs or VH CDRs corresponding toone or more of the human scFvs referred to in Table 1, or fragments orvariants thereof, and framework regions (and, optionally CDRs notderived from the scFvs in Table 1) from a human immunoglobulin molecule.In a preferred embodiment, an antibody of the present inventioncomprises a VH CDR3, VL CDR3, or both, corresponding to the same scFv,or different scFvs referred to in Table 1, or fragments or variantsthereof, and framework regions from a human immunoglobulin.

A chimeric antibody is a molecule in which different portions of theantibody are derived from different immunoglobulin molecules such asantibodies having a variable region derived from a human antibody and anon-human immunoglobulin constant region. Methods for producing chimericantibodies are known in the art. See e.g., Morrison, Science 229:1202(1985); Oi et al., BioTechniques 4:214 (1986); Gillies et al., J.Immunol Methods 125:191-202 (1989); U.S. Pat. Nos. 5,807,715; 4,816,567;and 4,816,397, which are incorporated herein by reference in theirentirety. Chimeric antibodies comprising one or more CDRs from humanspecies and framework regions from a non-human immunoglobulin molecule(e.g., framework regions from a canine or feline immunoglobulinmolecule) can be produced using a variety of techniques known in the artincluding, for example, CDR-grafting (EP 239,400; PCT publication WO91/09967; U.S. Pat. Nos. 5,225,539; 5,530,101; and 5,585,089), veneeringor resurfacing (EP 592,106; EP 519,596; Padlan, Molecular Immunology28(4/5):489-498 (1991); Studnicka et al., Protein Engineering7(6):805-814 (1994); Roguska et al., PNAS 91:969-973 (1994)), and chainshuffling (U.S. Pat. No. 5,565,332). In a preferred embodiment, chimericantibodies comprise a human CDR3 having an amino acid sequence of anyone of the VH CDR3s or VL CDR3s referred to in Table 1, or a variantthereof, and non-human framework regions or human framework regionsdifferent from those of the frameworks in the corresponding scFvdisclosed in Table 1. Often, framework residues in the framework regionswill be substituted with the corresponding residue from the CDR donorantibody to alter, preferably improve, antigen binding. These frameworksubstitutions are identified by methods well known in the art, e.g., bymodeling of the interactions of the CDR and framework residues toidentify framework residues important for antigen binding and sequencecomparison to identify unusual framework residues at particularpositions. (See, e.g., Queen et al., U.S. Pat. No. 5,585,089; Riechmannet al., Nature 332:323 (1988), which are incorporated herein byreference in their entireties.)

Further, the antibodies of the invention can, in turn, be utilized togenerate anti-idiotype antibodies that “mimic” B Lymphocyte Stimulatorpolypeptides using techniques well known to those skilled in the art.(See, e.g., Greenspan & Bona, FASEB J. 7(5):437-444 (1993); andNissinoff, J. Immunol. 147(8):2429-2438 (1991)). For example, antibodiesof the invention which bind to B Lymphocyte Stimulator and competitivelyinhibit the binding of B Lymphocyte Stimulator to its receptor (asdetermined by assays well known in the art such as, for example, thatdisclosed, infra) can be used to generate antiidiotypes that “mimic” a BLymphocyte Stimulator ligand/receptor-binding domain and, as aconsequence, bind to and neutralize B Lymphocyte Stimulator receptors(e.g., TACI—GenBank accession number AAC51790; BCMA—GenBank accessionnumber NP_(—)001183; and/or BAFF-R—GenBank accession numberNP_(—)443177). Such neutralizing anti-idiotypes (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants, such as Fab fragments of such anti-idiotypes) can be used intherapeutic regimens to neutralize B Lymphocyte Stimulator. For example,such anti-idiotypic antibodies can be used to bind B LymphocyteStimulator ligands/receptors, and thereby block B Lymphocyte Stimulatormediated biological activity. Alternatively, anti-idiotypes that “mimic”a B Lymphocyte Stimulator binding domain may bind to B LymphocyteStimulator receptor(s) and induce B Lymphocyte Stimulator receptormediated signalling (e.g., activation of nuclear factor of activated Tcells (NF-AT), nuclear factor-kappa B (NF-kappa B), and/or AP-1). Suchagonistic anti-idiotypes (including agonistic Fab fragments of theseanti-idiotypes) can be used in therapeutic regimens to induce or enhanceB Lymphocyte Stimulator receptor mediated signalling. For example, suchanti-idiotypic antibodies can be used to bind B Lymphocyte Stimulatorligands/receptors, and thereby stimulate B Lymphocyte Stimulatormediated biological activity (e.g., B cell proliferation and/orimmunoglobulin production).

Once an antibody molecule of the invention (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) has been chemically synthesized or recombinantlyexpressed, it may be purified by any method known in the art forpurification of an immunoglobulin molecule, or more generally, a proteinmolecule, such as, for example, by chromatography (e.g., ion exchange,affinity, particularly by affinity for the specific antigen afterProtein A, and sizing column chromatography), centrifugation,differential solubility, or by any other standard technique for thepurification of proteins. Further, the antibodies of the presentinvention may be fused to heterologous polypeptide sequences describedherein or otherwise known in the art, to facilitate purification.

Polynucleotides Encoding an Antibody

The invention provides polynucleotides comprising, or alternativelyconsisting of, a nucleotide sequence encoding an antibody of theinvention (including molecules comprising, or alternatively consistingof, antibody fragments or variants thereof). The invention alsoencompasses polynucleotides that hybridize under high stringency, oralternatively, under intermediate or lower stringency hybridizationconditions, e.g., as defined supra, to polynucleotides complementary tonucleic acids having a polynucleotide sequence that encodes an antibodyof the invention or a fragment or variant thereof.

The polynucleotides may be obtained, and the nucleotide sequence of thepolynucleotides determined, by any method known in the art. Since theamino acid sequences of the scFv antibodies and VH domains, VL domainsand CDRs thereof, are known (as described in Table 1), nucleotidesequences encoding these antibodies can be determined using methods wellknown in the art, i.e., the nucleotide codons known to encode theparticular amino acids are assembled in such a way to generate a nucleicacid that encodes the antibody, of the invention. Such a polynucleotideencoding the antibody may be assembled from chemically synthesizedoligonucleotides (e.g., as described in Kutmeier et al., BioTechniques17:242 (1994)), which, briefly, involves the synthesis of overlappingoligonucleotides containing portions of the sequence encoding theantibody, annealing and ligating of those oligonucleotides, and thenamplification of the ligated oligonucleotides by PCR.

Alternatively, a polynucleotide encoding an antibody (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) may be generated from nucleic acid from a suitablesource. If a clone containing a nucleic acid encoding a particularantibody is not available, but the sequence of the antibody molecule isknown, a nucleic acid encoding the immunoglobulin may be chemicallysynthesized or obtained from a suitable source (e.g., an antibody cDNAlibrary, or a cDNA library generated from, or nucleic acid, preferablypoly A+ RNA, isolated from, any tissue or cells expressing the antibody,such as hybridoma cells selected to express an antibody of theinvention) by PCR amplification using synthetic primers hybridizable tothe 3′ and 5′ ends of the sequence or by cloning using anoligonucleotide probe specific for the particular gene sequence toidentify, e.g., a cDNA clone from a cDNA library that encodes theantibody. Amplified nucleic acids generated by PCR may then be clonedinto replicable cloning vectors using any method well known in the art.

Once the nucleotide sequence of the antibody (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) is determined, the nucleotide sequence of the antibodymay be manipulated using methods well known in the art for themanipulation of nucleotide sequences, e.g., recombinant DNA techniques,site directed mutagenesis, PCR, etc. (see, for example, the techniquesdescribed in Sambrook et al., 1990, Molecular Cloning, A LaboratoryManual, 2d Ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.and Ausubel et al., eds., 1998, Current Protocols in Molecular Biology,John Wiley & Sons, NY, which are both incorporated by reference hereinin their entireties), to generate antibodies having a different aminoacid sequence, for example to create amino acid substitutions,deletions, and/or insertions.

In a specific embodiment, one or more of the VH and VL domains referredto in Table 1, or fragments or variants thereof, is inserted withinframework regions using recombinant DNA techniques known in the art. Ina specific embodiment, one, two, three, four, five, six, or more of theCDRs referred to in Table 1, or fragments or variants thereof, isinserted within framework regions using recombinant DNA techniques knownin the art. The framework regions may be naturally occurring orconsensus framework regions, and preferably human framework regions(see, e.g., Chothia et al., J. Mol. Biol. 278: 457-479 (1998) for alisting of human framework regions, the contents of which are herebyincorporated by reference in its entirety). Preferably, thepolynucleotides generated by the combination of the framework regionsand CDRs encode an antibody (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof)that specifically binds to B Lymphocyte Stimulator. Preferably, asdiscussed supra, polynucleotides encoding variants of antibodies orantibody fragments having one or more amino acid substitutions may bemade within the framework regions, and, preferably, the amino acidsubstitutions improve binding of the antibody to its antigen.Additionally, such methods may be used to make amino acid substitutionsor deletions of one or more variable region cysteine residuesparticipating in an intrachain disulfide bond to generate antibodymolecules, or antibody fragments or variants, lacking one or moreintrachain disulfide bonds. Other alterations to the polynucleotide areencompassed by the present invention and fall within the ordinary skillof the art.

Recombinant Expression of an Antibody

Recombinant expression of an antibody of the invention (including scFvsand other molecules comprising, or alternatively consisting of, antibodyfragments or variants thereof (e.g., a heavy or light chain of anantibody of the invention or a portion thereof or a single chainantibody of the invention)), requires construction of an expressionvector(s) containing a polynucleotide that encodes the antibody. Once apolynucleotide encoding an antibody molecule (e.g., a whole antibody, aheavy or light chain of an antibody, or portion thereof (preferably, butnot necessarily, containing the heavy or light chain variable domain)),of the invention has been obtained, the vector(s) for the production ofthe antibody molecule may be produced by recombinant DNA technologyusing techniques well known in the art. Thus, methods for preparing aprotein by expressing a polynucleotide containing an antibody encodingnucleotide sequence are described herein. Methods which are well knownto those skilled in the art can be used to construct expression vectorscontaining antibody coding sequences and appropriate transcriptional andtranslational control signals. These methods include, for example, invitro recombinant DNA techniques, synthetic techniques, and in vivogenetic recombination. The invention, thus, provides replicable vectorscomprising a nucleotide sequence encoding an antibody molecule of theinvention (e.g., a whole antibody, a heavy or light chain of anantibody, a heavy or light chain variable domain of an antibody, or aportion thereof, or a heavy or light chain CDR, a single chain Fv, orfragments or variants thereof), operably linked to a promoter. Suchvectors may include the nucleotide sequence encoding the constant regionof the antibody molecule (see, e.g., PCT Publication WO 86/05807; PCTPublication WO 89/01036; and U.S. Pat. No. 5,122,464, the contents ofeach of which are hereby incorporated by reference in its entirety) andthe variable domain of the antibody may be cloned into such a vector forexpression of the entire heavy chain, the entire light chain, or boththe entire heavy and light chains.

The expression vector(s) is(are) transferred to a host cell byconventional techniques and the transfected cells are then cultured byconventional techniques to produce an antibody of the invention. Thus,the invention includes host cells containing polynucleotide(s) encodingan antibody of the invention (e.g., whole antibody, a heavy or lightchain thereof, or portion thereof, or a single chain antibody of theinvention, or a fragment or variant thereof), operably linked to aheterologous promoter. In preferred embodiments, for the expression ofentire antibody molecules, vectors encoding both the heavy and lightchains may be co-expressed in the host cell for expression of the entireimmunoglobulin molecule, as detailed below.

A variety of host-expression vector systems may be utilized to expressthe antibody molecules of the invention. Such host-expression systemsrepresent vehicles by which the coding sequences of interest may beproduced and subsequently purified, but also represent cells which may,when transformed or transfected with the appropriate nucleotide codingsequences, express an antibody molecule of the invention in situ. Theseinclude, but are not limited to, microorganisms such as bacteria (e.g.,E. coli, B. subtilis) transformed with recombinant bacteriophage DNA,plasmid DNA or cosmid DNA expression vectors containing antibody codingsequences; yeast (e.g., Saccharomyces, Pichia) transformed withrecombinant yeast expression vectors containing antibody codingsequences; insect cell systems infected with recombinant virusexpression vectors (e.g., baculovirus) containing antibody codingsequences; plant cell systems infected with recombinant virus expressionvectors (e.g., cauliflower mosaic virus, CaMV; tobacco mosaic virus,TMV) or transformed with recombinant plasmid expression vectors (e.g.,Ti plasmid) containing antibody coding sequences; or mammalian cellsystems (e.g., COS, CHO, BHK, 293, 3T3 cells) harboring recombinantexpression constructs containing promoters derived from the genome ofmammalian cells (e.g., metallothionein promoter) or from mammalianviruses (e.g., the adenovirus late promoter; the vaccinia virus 7.5Kpromoter). Preferably, bacterial cells such as Escherichia coli, andmore preferably, eukaryotic cells, especially for the expression ofwhole recombinant antibody molecule, are used for the expression of arecombinant antibody molecule. For example, mammalian cells such asChinese hamster ovary cells (CHO), in conjunction with a vector such asthe major intermediate early gene promoter element from humancytomegalovirus is an effective expression system for antibodies(Foecking et al., Gene 45:101 (1986); Cockett et al., Bio/Technology 8:2(1990)).

In bacterial systems, a number of expression vectors may beadvantageously selected depending upon the use intended for the antibodymolecule being expressed. For example, when a large quantity of such aprotein is to be produced, for the generation of pharmaceuticalcompositions of an antibody molecule, vectors which direct theexpression of high levels of fusion protein products that are readilypurified may be desirable. Such vectors include, but are not limited to,the E. coli expression vector pUR278 (Ruther et al., EMBO 1. 2:1791(1983)), in which the antibody coding sequence may be ligatedindividually into the vector in frame with the lac Z coding region sothat a fusion protein is produced; pIN vectors (Inouye & Inouye, NucleicAcids Res. 13:3101-3109 (1985); Van Heeke & Schuster, J. Biol. Chem.24:5503-5509 (1989)); and the like. pGEX vectors may also be used toexpress foreign polypeptides as fusion proteins with glutathione5-transferase (GST). In general, such fusion proteins are soluble andcan easily be purified from lysed cells by adsorption and binding tomatrix glutathione agarose beads followed by elution in the presence offree glutathione. The pGEX vectors are designed to include thrombin orfactor Xa protease cleavage sites so that the cloned target gene productcan be released from the GST moiety.

In an insect system, Autographa californica nuclear polyhedrosis virus(AcNPV) may be used as a vector to express foreign genes. The virusgrows in Spodoptera frugiperda cells. Antibody coding sequences may becloned individually into non-essential regions (for example, thepolyhedrin gene) of the virus and placed under control of an AcNPVpromoter (for example, the polyhedrin promoter).

In mammalian host cells, a number of viral-based expression systems maybe utilized. In cases where an adenovirus is used as an expressionvector, the antibody coding sequence of interest may be ligated to anadenovirus transcription/translation control complex, e.g., the latepromoter and tripartite leader sequence. This chimeric gene may then beinserted in the adenovirus genome by in vitro or in vivo recombination.Insertion in a non-essential region of the viral genome (e.g., region E1or E3) will result in a recombinant virus that is viable and capable ofexpressing the antibody molecule in infected hosts (e.g., see Logan &Shenk, Proc. Natl. Acad. Sci. USA 8 1:355-359 (1984)). Specificinitiation signals may also be required for efficient translation ofinserted antibody coding sequences. These signals include the ATGinitiation codon and adjacent sequences. Furthermore, the initiationcodon must be in phase with the reading frame of the desired codingsequence to ensure translation of the entire insert. These exogenoustranslational control signals and initiation codons can be of a varietyof origins, both natural and synthetic. The efficiency of expression maybe enhanced by the inclusion of appropriate transcription enhancerelements, transcription terminators, etc. (see, e.g., Bittner et al.,Methods in Enzymol. 153:51-544 (1987)).

In addition, a host cell strain may be chosen which modulates theexpression of the inserted sequences, or modifies and processes the geneproduct in the specific fashion desired. Such modifications (e.g.,glycosylation) and processing (e.g., cleavage) of protein products maybe important for the function of the protein. Different host cells havecharacteristic and specific mechanisms for the post-translationalprocessing and modification of proteins and gene products. Appropriatecell lines or host systems can be chosen to ensure the correctmodification and processing of the foreign protein expressed. To thisend, eukaryotic host cells which possess the cellular machinery forproper processing of the primary transcript, glycosylation, andphosphorylation of the gene product may be used. Such mammalian hostcells include, but are not limited to, CHO, VERY, BHK, Hela, COS, NSO,MDCK, 293, 3T3, W138, and in particular, breast cancer cell lines suchas, for example, BT483, Hs578T, HTB2, BT2O and T47D, and normal mammarygland cell line such as, for example, CRL7O3O and HsS78Bst.

For long-term, high-yield production of recombinant proteins, stableexpression is preferred. For example, cell lines which stably expressthe antibody may be engineered. Rather than using expression vectorswhich contain viral origins of replication, host cells can betransformed with DNA controlled by appropriate expression controlelements (e.g., promoter, enhancer, sequences, transcriptionterminators, polyadenylation sites, etc.), and a selectable marker.Following the introduction of the foreign DNA, engineered cells may beallowed to grow for 1-2 days in an enriched media, and then are switchedto a selective media. The selectable marker in the recombinant plasmidconfers resistance to the selection and allows cells to stably integratethe plasmid into their chromosomes and grow to form foci which in turncan be cloned and expanded into cell lines. This method mayadvantageously be used to engineer cell lines which express the antibodymolecule. Such engineered cell lines may be particularly useful inscreening and evaluation of compositions that interact directly orindirectly with the antibody molecule.

A number of selection systems may be used, including but not limited to,the herpes simplex virus thymidine kinase (Wigler et al., Cell 11:223(1977)), hypoxanthineguanine phosphoribosyltransferase (Szybalska &Szybalski, Proc. Natl. Acad. Sci. USA 48:202 (1992)), and adeninephosphoribosyltransferase (Lowy et al., Cell 22:8 17 (1980)) genes canbe employed in tk-, hgprt- or aprt-cells, respectively. Also,antimetabolite resistance can be used as the basis of selection for thefollowing genes: dhfr, which confers resistance to methotrexate (Wigleret al., Natl. Acad. Sci. USA 77:357 (1980); O'Hare et al., Proc. Natl.Acad. Sci. USA 78:1527 (1981)); gpt, which confers resistance tomycophenolic acid (Mulligan & Berg, Proc. Natl. Acad. Sci. USA 78:2072(1981)); neo, which confers resistance to the aminoglycoside G-418(Clinical Pharmacy 12:488-505; Wu and Wu, Biotherapy 3:87-95 (1991);Tolstoshev, Ann. Rev. Pharmacol. Toxicol. 32:573-596 (1993); Mulligan,Science 260:926-932 (1993); and Morgan and Anderson, Ann. Rev. Biochem.62: 191-217 (1993); TIB TECH 11(5):155-215 (May, 1993)); and hygro,which confers resistance to hygromycin (Santerre et al., Gene 30:147(1984)). Methods commonly known in the art of recombinant DNA technologymay be routinely applied to select the desired recombinant clone, andsuch methods are described, for example, in Ausubel et al. (eds.),Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993);Kriegler, Gene Transfer and Expression, A Laboratory Manual, StocktonPress, NY (1990); and in Chapters 12 and 13, Dracopoli et al. (eds),Current Protocols in Human Genetics, John Wiley & Sons, NY (1994);Colberre-Garapin et al., J. Mol. Biol. 150:1 (1981), which areincorporated by reference herein in their entireties.

The expression levels of an antibody molecule can be increased by vectoramplification (for a review, see Bebbington and Hentschel, The use ofvectors based on gene amplification for the expression of cloned genesin mammalian cells in DNA cloning, Vol. 3. (Academic Press, New York,1987)). When a marker in the vector system expressing antibody isamplifiable, increase in the level of inhibitor present in culture ofhost cell will increase the number of copies of the marker gene. Sincethe amplified region is associated with the coding sequence of theantibody, production of the antibody will also increase (Crouse et al.,Mol. Cell. Biol. 3:257 (1983)).

The host cell may be co-transfected with two expression vectors of theinvention, the first vector encoding a heavy chain derived polypeptideand the second vector encoding a light chain derived polypeptide. Thetwo vectors may contain identical selectable markers which enable equalexpression of heavy and light chain polypeptides. Alternatively, asingle vector may be used which encodes, and is capable of expressing,both heavy and light chain polypeptides. In such situations, the lightchain is preferably placed before the heavy chain to avoid an excess oftoxic free heavy chain (Proudfoot, Nature 322:52 (1986); Kohler, Proc.Natl. Acad. Sci. USA 77:2 197 (1980)). The coding sequences for theheavy and light chains may comprise cDNA or genomic DNA.

Once an antibody molecule of the invention has been produced byrecombinant expression, it may be purified by any method known in theart for purification of an immunoglobulin molecule, or more generally,for purification of a protein, for example, by chromatography (e.g., ionexchange, affinity, particularly by affinity for the specific antigenafter Protein A, and sizing column chromatography), centrifugation,differential solubility, or by any other standard technique for thepurification of proteins. Further, the antibodies of the presentinvention may be fused to heterologous polypeptide sequences describedherein or otherwise known in the art to facilitate purification.

Antibodies of the present invention include naturally purified products,products of chemical synthetic procedures, and products produced byrecombinant techniques from a prokaryotic or eukaryotic host, including,for example, bacterial, yeast, higher plant, insect and mammalian cells.Depending upon the host employed in a recombinant production procedure,the antibodies of the present invention may be glycosylated or may benon-glycosylated. In addition, antibodies of the invention may alsoinclude an initial modified methionine residue, in some cases as aresult of host-mediated processes.

Antibodies of the invention can be chemically synthesized usingtechniques known in the art (e.g., see Creighton, 1983, Proteins:Structures and Molecular Principles, W.H. Freeman & Co., N.Y., andHunkapiller, M., et al., 1984, Nature 310:105-111). For example, apeptide corresponding to a fragment of an antibody of the invention canbe synthesized by use of a peptide synthesizer. Furthermore, if desired,nonclassical amino acids or chemical amino acid analogs can beintroduced as a substitution or addition into the antibody polypeptidesequence. Non-classical amino acids include, but are not limited to, tothe D-isomers of the common amino acids, 2,4-diaminobutyric acid,a-amino isobutyric acid, 4-aminobutyric acid, Abu, 2-amino butyric acid,g-Abu, e-Ahx, 6-amino hexanoic acid, Aib, 2-amino isobutyric acid,3-amino propionic acid, ornithine, norleucine, norvaline,hydroxyproline, sarcosine, citrulline, homocitrulline, cysteic acid,t-butylglycine, t-butylalanine, phenylglycine, cyclohexylalanine,b-alanine, fluoro-amino acids, designer amino acids such as b-methylamino acids, Ca-methyl amino acids, Na-methyl amino acids, and aminoacid analogs in general. Furthermore, the amino acid can be D(dextrorotary) or L (levorotary).

The invention encompasses antibodies which are differentially modifiedduring or after translation, e.g., by glycosylation, acetylation,phosphorylation, amidation, derivatization by known protecting/blockinggroups, proteolytic cleavage, linkage to an antibody molecule or othercellular ligand, etc. Any of numerous chemical modifications may becarried out by known techniques, including but not limited, to specificchemical cleavage by cyanogen bromide, trypsin, chymotrypsin, papain, V8protease, NaBH₄, acetylation, formylation, oxidation, reduction,metabolic synthesis in the presence of tunicamycin, etc.

Additional post-translational modifications encompassed by the inventioninclude, for example, e.g., N-linked or O-linked carbohydrate chains,processing of N-terminal or C-terminal ends), attachment of chemicalmoieties to the amino acid backbone, chemical modifications of N-linkedor O-linked carbohydrate chains, and addition or deletion of anN-terminal methionine residue as a result of procaryotic host cellexpression. The polypeptides may also be modified with a detectablelabel, such as an enzymatic, fluorescent, radioisotopic or affinitylabel to allow for detection and isolation of the antibody.

Examples of suitable enzymes include horseradish peroxidase, alkalinephosphatase, beta-galactosidase, glucose oxidase oracetylcholinesterase; examples of suitable prosthetic group complexesinclude streptavidin/biotin and avidin/biotin; examples of suitablefluorescent materials include biotin, umbelliferone, fluorescein,fluorescein isothiocyanate, rhodamine, dichlorotriazinylaminefluorescein, dansyl chloride or phycoerythrin; an example of aluminescent material includes luminol; examples of bioluminescentmaterials include luciferase, luciferin, and aequorin; and examples ofsuitable radioactive material include a radioactive metal ion, e.g.,alpha-emitters such as, for example, ²¹³Bi, or other radioisotopes suchas, for example, iodine (¹³¹I, ¹²⁵I, ¹²³I, ¹²¹I), carbon (¹⁴C), sulfur(³⁵S), tritium (³H), indium (^(115m)In, ^(113m)In, ¹¹²In, ¹¹¹In), andtechnetium (⁹⁹Tc, ^(99m)Tc), thallium (²⁰¹Ti), gallium (⁶⁸Ga, ⁶⁷Ga),palladium (¹⁰³Pd), molybdenum (⁹⁹Mo), xenon (¹³³Xe), fluorine (¹⁸F),¹⁵³Sm, ¹⁷⁷Lu, ¹⁵⁹Gd, ¹⁴⁹Pm, ¹⁴⁰La, ¹⁷⁵Yb, ¹⁶⁶HO, ⁹⁰Y, ⁴⁷Sc, ¹⁸⁶Re, ¹⁸⁸_(Re,) ¹⁴²Pr, ¹⁰⁵Rh, ⁹⁷Ru, ⁶⁸Ge, ⁵⁷Co, ⁶⁵Zn, ⁸⁵Sr, ³²P, ¹⁵³Gd, ¹⁶⁹Yb,⁵¹Cr, ⁵⁴Mn, ⁷⁵Se, ¹¹³Sn, and ¹¹⁷Tin.

In specific embodiments, antibodies of the invention may be labeled withEuropium. For example, antibodies of the invention may be labelled withEuropium using the DELFIA Eu-labeling kit (catalog #1244-302, PerkinElmer Life Sciences, Boston, Mass.) following manufacturer'sinstructions.

In specific embodiments, antibodies of the invention are attached tomacrocyclic chelators useful for conjugating radiometal ions, includingbut not limited to, ¹¹¹In, ¹⁷⁷Lu, ⁹⁰Y, ¹⁶⁶Ho, and ¹⁵³Sm, to antibodies.In a preferred embodiment, the radiometal ion associated with themacrocyclic chelators attached to antibodies of the invention is ¹¹¹In.In another preferred embodiment, the radiometal ion associated with themacrocyclic chelator attached to antibodies of the invention is ⁹⁰Y. Inspecific embodiments, the macrocyclic chelator is1,4,7,10-tetraazacyclododecane-N,N′,N″,N′″-tetraacetic acid (DOTA). Inspecific embodiments, the macrocyclic chelator isα-(5-isothiocyanato-2-methoxyphenyl)-1,4,7,10-tetraaza-cyclododecane-1,4,7,10-tetraaceticacid. In other specific embodiments, the DOTA is attached to theantibody of the invention via a linker molecule. Examples of linkermolecules useful for conjugating DOTA to an antibody are commonly knownin the art—see, for example, DeNardo et al., Clin Cancer Res.4(10):2483-90, 1998; Peterson et al., Bioconjug. Chem. 10(4):553-7,1999; and Zimmerman et al, Nucl. Med. Biol. 26(8):943-50, 1999 which arehereby incorporated by reference in their entirety. In addition, U.S.Pat. Nos. 5,652,361 and 5,756,065, which disclose chelating agents thatmay be conjugated to antibodies, and methods for making and using them,are hereby incorporated by reference in their entireties.

In one embodiment, antibodies of the invention are labeled with biotin.In other related embodiments, biotinylated antibodies of the inventionmay be used, for example, as an imaging agent or as a means ofidentifying one or more B Lymphocyte Stimulator receptor(s) or othercoreceptor or ligand molecules.

Also provided by the invention are chemically modified derivatives ofantibodies of the invention which may provide additional advantages suchas increased solubility, stability and in vivo or in vitro circulatingtime of the polypeptide, or decreased immunogenicity (see U.S. Pat. No.4,179,337). The chemical moieties for derivitization may be selectedfrom water soluble polymers such as polyethylene glycol, ethyleneglycol/propylene glycol copolymers, carboxymethylcellulose, dextran,polyvinyl alcohol and the like. The polypeptides may be modified atrandom positions within the molecule, or at predetermined positionswithin the molecule and may include one, two, three or more attachedchemical moieties.

The polymer may be of any molecular weight, and may be branched orunbranched. For polyethylene glycol, the preferred molecular weight isbetween about 1 kDa and about 100 kDa (the term “about” indicating thatin preparations of polyethylene glycol, some molecules will weigh more,some less, than the stated molecular weight) for ease in handling andmanufacturing. Other sizes may be used, depending on the desiredtherapeutic profile (e.g., the duration of sustained release desired,the effects, if any on biological activity, the ease in handling, thedegree or lack of antigenicity and other known effects of thepolyethylene glycol to a therapeutic protein or analog). For example,the polyethylene glycol may have an average molecular weight of about200, 500, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500,6000, 6500, 7000, 7500, 8000, 8500, 9000, 9500, 10,000, 10,500, 11,000,11,500, 12,000, 12,500, 13,000, 13,500, 14,000, 14,500, 15,000, 15,500,16,000, 16,500, 17,000, 17,500, 18,000, 18,500, 19,000, 19,500, 20,000,25,000, 30,000, 35,000, 40,000, 50,000, 55,000, 60,000, 65,000, 70,000,75,000, 80,000, 85,000, 90,000, 95,000, or 100,000 kDa.

As noted above, the polyethylene glycol may have a branched structure.Branched polyethylene glycols are described, for example, in U.S. Pat.No. 5,643,575; Morpurgo et al., Appl. Biochem. Biotechnol. 56:59-72(1996); Vorobjev et al., Nucleosides Nucleotides 18:2745-2750 (1999);and Caliceti et al., Bioconjug. Chem. 10:638-646 (1999), the disclosuresof each of which are incorporated herein by reference.

The polyethylene glycol molecules (or other chemical moieties) should beattached to the protein with consideration of effects on functional orantigenic domains of the antibody. There are a number of attachmentmethods available to those skilled in the art, e.g., EP 0 401 384,herein incorporated by reference (coupling PEG to G-CSF), see also Maliket al., Exp. Hematol. 20:1028-1035 (1992) (reporting pegylation ofGM-CSF using tresyl chloride). For example, polyethylene glycol may becovalently bound through amino acid residues via a reactive group, suchas, a free amino or carboxyl group. Reactive groups are those to whichan activated polyethylene glycol molecule may be bound. The amino acidresidues having a free amino group may include, for example, lysineresidues and the N-terminal amino acid residues; those having a freecarboxyl group may include aspartic acid residues, glutamic acidresidues, and the C-terminal amino acid residue. Sulfhydryl groups mayalso be used as a reactive group for attaching the polyethylene glycolmolecules. Preferred for therapeutic purposes is attachment at an aminogroup, such as attachment at the N-terminus or lysine group.

As suggested above, polyethylene glycol may be attached to proteins,e.g., antibodies, via linkage to any of a number of amino acid residues.For example, polyethylene glycol can be linked to a proteins viacovalent bonds to lysine, histidine, aspartic acid, glutamic acid, orcysteine residues. One or more reaction chemistries may be employed toattach polyethylene glycol to specific amino acid residues (e.g.,lysine, histidine, aspartic acid, glutamic acid, or cysteine) of theantibody or to more than one type of amino acid residue (e.g., lysine,histidine, aspartic acid, glutamic acid, cysteine and combinationsthereof) of the antibody.

One may specifically desire antibodies chemically modified at theN-terminus of either the heavy chain or the light chain or both. Usingpolyethylene glycol as an illustration, one may select from a variety ofpolyethylene glycol molecules (by molecular weight, branching, etc.),the proportion of polyethylene glycol molecules to protein (or peptide)molecules in the reaction mix, the type of pegylation reaction to beperformed, and the method of obtaining the selected N-terminallypegylated protein. The method of obtaining the N-terminally pegylatedpreparation (i.e., separating this moiety from other monopegylatedmoieties if necessary) may be by purification of the N-terminallypegylated material from a population of pegylated protein molecules.Selective chemical modification at the N-terminus may be accomplished byreductive alkylation which exploits differential reactivity of differenttypes of primary amino groups (lysine versus the N-terminal) availablefor derivatization in a particular antibody, e.g., a heavy chain oralight chain. Under the appropriate reaction conditions, substantiallyselective derivatization of the protein at the N-terminus with acarbonyl group containing polymer is achieved.

As indicated above, pegylation of the proteins of the invention may beaccomplished by any number of means. For example, polyethylene glycolmay be attached to the protein either directly or by an interveninglinker. Linkerless systems for attaching polyethylene glycol to proteinsare described in Delgado et al., Crit. Rev. Thera. Drug Carrier Sys.9:249-304 (1992); Francis et al., Intern. J. of Hematol. 68:1-18 (1998);U.S. Pat. No. 4,002,531; U.S. Pat. No. 5,349,052; WO 95/06058; and WO98/32466, the disclosures of each of which are incorporated herein byreference.

One system for attaching polyethylene glycol directly to amino acidresidues of proteins without an intervening linker employs tresylatedMPEG, which is produced by the modification of monmethoxy polyethyleneglycol (MPEG) using tresylchloride (ClSO₂CH₂CF₃). Upon reaction ofantibody with tresylated MPEG, polyethylene glycol is directly attachedto amine groups of the antibody. Thus, the invention includesantibody-polyethylene glycol conjugates produced by reacting proteins ofthe invention with a polyethylene glycol molecule having a2,2,2-trifluoreothane sulphonyl group.

Polyethylene glycol can also be attached to antibodies using a number ofdifferent intervening linkers. For example, U.S. Pat. No. 5,612,460, theentire disclosure of which is incorporated herein by reference,discloses urethane linkers for connecting polyethylene glycol toproteins. Protein-polyethylene glycol conjugates wherein thepolyethylene glycol is attached to the antibody by a linker can also beproduced by reaction of antibodies with compounds such asMPEG-succinimidylsuccinate, MPEG activated with1,1′-carbonyldiimidazole, MPEG-2,4,5-trichloropenylcarbonate,MPEG-p-nitrophenolcarbonate, and various MPEG-succinate derivatives. Anumber additional polyethylene glycol derivatives and reactionchemistries for attaching polyethylene glycol to proteins are describedin WO 98/32466, the entire disclosure of which is incorporated herein byreference. Pegylated protein products produced using the reactionchemistries set out herein are included within the scope of theinvention.

The number of polyethylene glycol moieties attached to each antibody ofthe invention (i.e., the degree of substitution) may also vary. Forexample, the pegylated antibodies of the invention may be linked, onaverage, to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 17, 20, or morepolyethylene glycol molecules. Similarly, the average degree ofsubstitution within ranges such as 1-3, 2-4, 3-5, 4-6, 5-7, 6-8, 7-9,8-10, 9-11, 10-12, 11-13, 12-14, 13-15, 14-16, 15-17, 16-18, 17-19, or18-20 polyethylene glycol moieties per antibody molecule. Methods fordetermining the degree of substitution are discussed, for example, inDelgado et al., Crit. Rev. Thera. Drug Carrier Sys. 9:249-304 (1992).

Antibody Characterization

Antibodies of the present invention (including scFvs and other moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) may be characterized in a variety of ways. Inparticular, antibodies and related molecules of the invention may beassayed for the ability to immunospecifically bind to B LymphocyteStimulator or a fragment of B Lymphocyte Stimulator (e.g., to thesoluble form or the membrane-bound form of B Lymphocyte Stimulator)using techniques described herein or routinely modifying techniquesknown in the art. B Lymphocyte Stimulator or B Lymphocyte Stimulatorfragments that may be immunospecifically bound by the compositions ofthe invention include, but are not limited to, human B LymphocyteStimulator (SEQ ID NOS:3228 and/or 3229) or B Lymphocyte Stimulatorexpressed on human monocytes; murine B Lymphocyte Stimulator (SEQ IDNOS:3230 and/or 3231) or B Lymphocyte Stimulator expressed on murinemonocytes; rat B Lymphocyte Stimulator (either the soluble forms asgiven in SEQ ID NOS:3232, 3233, 3234 and/or 3235 or in a membraneassociated form, e.g., on the surface of rat monocytes); or monkey BLymphocyte Stimulator (e.g., the monkey B Lymphocyte Stimulatorpolypeptides of SEQ ID NOS:3236 and/or 3237, the soluble form of monkeyB Lymphocyte Stimulator, or B Lymphocyte Stimulator expressed on monkeymonocytes) or fragments thereof. Preferably compositions of theinvention bind human B Lymphocyte Stimulator (SEQ ID NOS:3228 and/or3229) or fragments thereof. Assays for the ability of the antibodies ofthe invention to immunospecifically bind B Lymphocyte Stimulator or afragment of B Lymphocyte Stimulator may be performed in solution (e.g.,Houghten, Bio/Techniques 13:412-421(1992)), on beads (e.g., Lam, Nature354:82-84 (1991)), on chips (e.g., Fodor, Nature 364:555-556 (1993)), onbacteria (e.g., U.S. Pat. No. 5,223,409), on spores (e.g., Pat. Nos.5,571,698; 5,403,484; and 5,223,409), on plasmids (e.g., Cull et al.,Proc. Natl. Acad. Sci. USA 89:1865-1869 (1992)) or on phage (e.g., Scottand Smith, Science 249:386-390 (1990); Devlin, Science 249:404-406(1990); Cwirla et al., Proc. Natl. Acad. Sci. USA 87:6378-6382 (1990);and Felici, J. Mol. Biol. 222:301-310 (1991)) (each of these referencesis incorporated herein in its entirety by reference). Antibodies thathave been identified to immunospecifically bind to B LymphocyteStimulator or a fragment of B Lymphocyte Stimulator can then be assayedfor their specificity and affinity for B Lymphocyte Stimulator or afragment of B Lymphocyte Stimulator using or routinely modifyingtechniques described herein or otherwise known in the art.

The antibodies of the invention may be assayed for immunospecificbinding to B Lymphocyte Stimulator and cross-reactivity with otherantigens by any method known in the art. In particular, the ability ofan antibody to immunospecifically bind to the soluble form ormembrane-bound form of B Lymphocyte Stimulator and the specificity ofthe antibody, fragment, or variant for B Lymphocyte Stimulatorpolypeptide from a particular species (e.g., murine, monkey or human,preferably human) may be determined using or routinely modifyingtechniques described herein or otherwise known in art.

Immunoassays which can be used to analyze immunospecific binding andcross-reactivity include, but are not limited to, competitive andnon-competitive assay systems using techniques such as western blots,radioimmunoassays, ELISA (enzyme linked immunosorbent assay), “sandwich”immunoassays, immunoprecipitation assays, precipitant reactions, geldiffusion precipitant reactions, immunodiffusion assays, agglutinationassays, complement-fixation assays, immunoradiometric assays,fluorescent immunoassays, and protein A immunoassays, to name but a few.Such assays are routine and well known in the art (see, e.g., Ausubel etal, eds, 1994, Current Protocols in Molecular Biology, Vol. 1, JohnWiley & Sons, Inc., New York, which is incorporated by reference hereinin its entirety). Exemplary immunoassays are described briefly below(but are not intended by way of limitation).

Immunoprecipitation protocols generally comprise lysing a population ofcells in a lysis buffer such as RIPA buffer (1% NP-40 or Triton X-100,1% sodium deoxycholate, 0.1% SDS, 0.15 M NaCl, 0.01 M sodium phosphateat pH 7.2, 1% Trasylol) supplemented with protein phosphatase and/orprotease inhibitors (e.g., EDTA, PMSF, aprotinin, sodium vanadate),adding the antibody of interest to the cell lysate, incubating for aperiod of time (e.g., 1 to 4 hours) at 40 degrees C., adding protein Aand/or protein G sepharose beads to the cell lysate, incubating forabout an hour or more at 40 degrees C., washing the beads in lysisbuffer and resuspending the beads in SDS/sample buffer. The ability ofthe antibody of interest to immunoprecipitate a particular antigen canbe assessed by, e.g., western blot analysis. One of skill in the artwould be knowledgeable as to the parameters that can be modified toincrease the binding of the antibody to an antigen and decrease thebackground (e.g., pre-clearing the cell lysate with sepharose beads).For further discussion regarding immunoprecipitation protocols see,e.g., Ausubel et al, eds, 1994, Current Protocols in Molecular Biology,Vol. 1, John Wiley & Sons, Inc., New York at 10.16.1.

Western blot analysis generally comprises preparing protein samples,electrophoresis of the protein samples in a polyacrylamide gel (e.g.,8%-20% SDS-PAGE depending on the molecular weight of the antigen),transferring the protein sample from the polyacrylamide gel to amembrane such as nitrocellulose, PVDF or nylon, blocking the membrane inblocking solution (e.g., PBS with 3% BSA or non-fat milk), washing themembrane in washing buffer (e.g., PBS-Tween 20), blocking the membranewith primary antibody (the antibody of interest) diluted in blockingbuffer, washing the membrane in washing buffer, blocking the membranewith a secondary antibody (which recognizes the primary antibody, e.g.,an anti-human antibody) conjugated to an enzymatic substrate (e.g.,horseradish peroxidase or alkaline phosphatase) or radioactive molecule(e.g., ³²P or ¹²⁵I) diluted in blocking buffer, washing the membrane inwash buffer, and detecting the presence of the antigen. One of skill inthe art would be knowledgeable as to the parameters that can be modifiedto increase the signal detected and to reduce the background noise. Forfurther discussion regarding western blot protocols see, e.g., Ausubelet al, eds, 1994, Current Protocols in Molecular Biology, Vol. 1, JohnWiley & Sons, Inc., New York at 10.8.1.

ELISAs comprise preparing antigen, coating the well of a 96-wellmicrotiter plate with the antigen, washing away antigen that did notbind the wells, adding the antibody of interest conjugated to adetectable compound such as an enzymatic substrate (e.g., horseradishperoxidase or alkaline phosphatase) to the wells and incubating for aperiod of time, washing away unbound antibodies or non-specificallybound antibodies, and detecting the presence of the antibodiesspecifically bound to the antigen coating the well. In ELISAs theantibody of interest does not have to be conjugated to a detectablecompound; instead, a second antibody (which recognizes the antibody ofinterest) conjugated to a detectable compound may be added to the well.Further, instead of coating the well with the antigen, the antibody maybe coated to the well. In this case, the detectable molecule could bethe antigen conjugated to a detectable compound such as an enzymaticsubstrate (e.g., horseradish peroxidase or alkaline phosphatase). One ofskill in the art would be knowledgeable as to the parameters that can bemodified to increase the signal detected as well as other variations ofELISAs known in the art. For further discussion regarding ELISAs see,e.g., Ausubel et al, eds, 1994, Current Protocols in Molecular Biology,Vol. 1, John Wiley & Sons, Inc., New York at 11.2.1.

The binding affinity of an antibody (including an scFv or other moleculecomprising, or alternatively consisting of, antibody fragments orvariants thereof) to an antigen and the off-rate of an antibody-antigeninteraction can be determined by competitive binding assays. One exampleof a competitive binding assay is a radioimmunoassay comprising theincubation of labeled antigen (e.g., ³H or ¹²⁵I) with the antibody ofinterest in the presence of increasing amounts of unlabeled antigen, andthe detection of the antibody bound to the labeled antigen. The affinityof the antibody of the present invention for B Lymphocyte Stimulator andthe binding off-rates can be determined from the data by Scatchard plotanalysis. Competition with a second antibody can also be determinedusing radioimmunoassays. In this case, B Lymphocyte Stimulator isincubated with an antibody of the present invention conjugated to alabeled compound (e.g., ³H or ¹²⁵I) in the presence of increasingamounts of an unlabeled second anti-B Lymphocyte Stimulator antibody.

In a preferred embodiment, BIAcore kinetic analysis is used to determinethe binding on and off rates of antibodies (including an scFv or othermolecule comprising, or alternatively consisting of, antibody fragmentsor variants thereof) to B Lymphocyte Stimulator, or fragments of BLymphocyte Stimulator. BIAcore kinetic analysis comprises analyzing thebinding and dissociation of B Lymphocyte Stimulator from chips withimmobilized antibodies on their surface as described in detail inExamples 6, 12, 17 and 18, infra.

The antibodies of the invention (including scFvs and other moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) can also be assayed for their ability to inhibit,increase, or not significantly alter, the binding of B LymphocyteStimulator to a B Lymphocyte Stimulator receptor (e.g., TACI—GenBankaccession number AAC51790; BCMA—GenBank accession number NP_(—)001183;and/or BAFF-R—GenBank accession number NP_(—)443177) using techniquesknown to those of skill in the art. For example, cells expressing areceptor for B Lymphocyte Stimulator (e.g., IM9, REH, ARH-77cells,Namalwa, and RPMI-8226 B cell tumor lines as wells as peripheral CD20+ Bcells) can be contacted with B Lymphocyte Stimulator in the presence orabsence of an antibody, and the ability of the antibody to inhibit,increase, or not significantly alter, B Lymphocyte Stimulator binding tothe cells can be measured. B Lymphocyte Stimulator binding to cells canbe measured by, for example, flow cytometry or a scintillation assay. BLymphocyte Stimulator or the antibody can be labeled with a detectablecompound such as a radioactive label (e.g., ³²P, ³⁵S, and ¹²⁵I) or afluorescent label (e.g., fluorescein isothiocyanate, rhodamine,phycoerythrin, phycocyanin, allophycocyanin, o-phthaldehyde andfluorescamine) to enable detection of an interaction between BLymphocyte Stimulator and a B Lymphocyte Stimulator receptor and/or BLymphocyte Stimulator and an antibody of the invention. Alternatively,the ability of antibodies of the invention to inhibit, increase, or notsignificantly alter, B Lymphocyte Stimulator binding to a B LymphocyteStimulator receptor can be determined in cell-free assays. For example,native or recombinant B Lymphocyte Stimulator (e.g., that having theamino acid sequence of amino acids 134-285 of SEQ ID NO:3228) or afragment thereof can be contacted with an antibody and the ability ofthe antibody to inhibit, increase, or not significantly alter, BLymphocyte Stimulator from binding to a B Lymphocyte Stimulator receptorcan be determined Preferably, the antibody is immobilized on a solidsupport and B Lymphocyte Stimulator or a B Lymphocyte Stimulatorfragment is labeled with a detectable compound. Alternatively, BLymphocyte Stimulator or a B Lymphocyte Stimulator fragment isimmobilized on a solid support and the antibody is labeled with adetectable compound. B Lymphocyte Stimulator may be partially orcompletely purified (e.g., partially or completely free of otherpolypeptides) or part of a cell lysate. Further, the B LymphocyteStimulator polypeptide may be a fusion protein comprising B LymphocyteStimulator or a biologically active portion thereof and a domain such asan Immunoglobulin Fc or glutathionine-S-transferase. For example, aminoacid residues 1-154 of TACI (GenBank accession number AAC51790), aminoacids 1-48 of BCMA (GenBank accession number NP_(—)001183) or aminoacids 1-81 of BAFF-R (GenBank Accession Number NP_(—)443177) may befused to the Fc region of an IgG molecule and used in a cell free assayto determine the ability of antibodies of the invention to inhibit,increase, or not significantly alter, B Lymphocyte Stimulator binding toa B Lymphocyte Stimulator receptor. Alternatively, B LymphocyteStimulator can be biotinylated using techniques well known to those ofskill in the art (e.g., biotinylation kit, Pierce Chemicals; Rockford,Ill.).

The antibodies of the invention (including scFvs or other moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), can also be assayed for their ability to inhibit,stimulate, or not significantly alter, B Lymphocyte Stimulator-inducedB-cell proliferation using techniques known to those of skill in theart. For example, B-cell proliferation can be assayed by ³H-thymidineincorporation assays and trypan blue cell counts (see, e.g., Moore etal., Science 285: 260-263 (1999)). Further, the antibodies of theinvention, or fragments or variants thereof, can be assayed for theirability to block, stimulate, or not significantly alter, B LymphocyteStimulator-induced activation of cellular signaling molecules andtranscription factors such as calcium-modulator and cyclophilin ligand(“CAML”), calcineurin, nuclear factor of activated T cells transcriptionfactor (“NF-AT”), nuclear factor-kappa B (“NF-kappa B”), and AP-1 usingtechniques known to those of skill in the art (see, e.g., von Bulow andBram, Science 278:138-141(1997)). For example, NF-AT activity can bedetermined by electromobility gel shift assays, by detecting theexpression of a protein known to be regulated by NF-AT (e.g., IL-2expression), by detecting the induction of a reporter gene (e.g., anNF-AT regulatory element operably linked to a nucleic acid encoding adetectable marker such as luciferase, beta-galactosidase orchloramphenicol acetyltransferase (CAT)), or by detecting a cellularresponse (e.g., cellular differentiation, or cell proliferation).

The antibodies of the invention, or fragments or variants thereof canalso be assayed for their ability to neutralize, enhance, or notsignificantly alter, B Lymphocyte Stimulator activity. For example,antibodies or fragments or variants thereof, may be routinely tested fortheir ability to inhibit B Lymphocyte Stimulator from binding to cellsexpressing the receptor for B Lymphocyte Stimulator (see Example 3,infra).

Selection and Screening for Antibodies that Immunospecifically Bind toSoluble B Lymphocyte Stimulator

Antibodies of the invention (including scFvs and other moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) may be screened in a variety of assays to identifythose antibodies that immunospecifically bind to the soluble form of BLymphocyte Stimulator. In one particular assay, antibodies that bind tothe biotinylated soluble form of B Lymphocyte Stimulator in solution arecaptured on streptavidin coated magnetic beads. This assay may berelatively applied to identify antibodies of the invention thatneutralize and/or bind to B Lymphocyte Stimulator. Additionally,antibodies may be assayed in neutralization assays described herein orotherwise known in the art (see Example 3, infra). For example,antibodies may be tested for their ability to inhibit soluble BLymphocyte Stimulator (e.g., biotinylated B Lymphocyte Stimulator) frombinding to IM9 cells. In this assay, labeled soluble B LymphocyteStimulator (e.g., biotinylated B Lymphocyte Stimulator) is incubatedwith candidate anti-B Lymphocyte Stimulator antibodies to allow for theformation of B Lymphocyte Stimulator -anti-B Lymphocyte Stimulatorantibody complexes. Following incubation, an aliquot of the B LymphocyteStimulator-anti-B Lymphocyte Stimulator antibody sample is added to IM9cells. The binding of soluble B Lymphocyte Stimulator may be determinedusing techniques known in the art. For example, the binding ofbiotinylated B Lymphocyte Stimulator to IM9 cells may be detected usinga fluorimeter following the addition of streptavidin-delfia.Biotinylated B Lymphocyte Stimulator, if it is not bound by antibodiesthat neutralize B Lymphocyte Stimulator, binds to the cells is detected.Thus, an antibody that decreases the amount of bio-B LymphocyteStimulator that binds to IM-9 cells (relative to a control sample inwhich the B Lymphocyte Stimulator had been preincubated with anirrelevant antibody or no antibody at all) is identified as one thatbinds to and neutralizes the soluble form of B Lymphocyte Stimulator. Inanother assay, antibodies are screened using ELISAs for those antibodiesthat bind to biotinylated soluble B Lymphocyte Stimulator, but do notbind membrane-bound B Lymphocyte Stimulator, such as, for example, BLymphocyte Stimulator on membranes from U937 cells (see Examples 2 and9, infra). In these assays, soluble B Lymphocyte Stimulator (e.g.,biotinylated B Lymphocyte Stimulator) and membrane-bound B LymphocyteStimulator (e.g., on U937 membranes) are incubated in separate sampleswith the same antibodies and those antibodies that bind to the soluble BLymphocyte Stimulator (biotinylated B Lymphocyte Stimulator), but notmembrane-bound B Lymphocyte Stimulator (e.g., on U937 membranes) arecaptured and identified.

Antibodies of the invention (including scFvs and other moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) may be tested to identify those antibodies that do notcross-react with APRIL, endokine-alpha, VEGI, TRAIL, TNF-alpha,TNF-beta, Fas-L, LIGHT, and PBS (see Example 4, infra). Antibodies mayalso be tested for their affinity for B Lymphocyte Stimulator using, forexample, BIAcore analysis (see Examples 6, 12, 17 and 18 infra).Antibodies may also be tested for their ability to stimulate, inhibit,or not alter, B Lymphocyte Stimulator-induced immunoglobulin productionand/or B-cell proliferation using techniques known to those of skill inthe art. For example, human B-cells, B Lymphocyte Stimulator andantibodies may be incubated together in 96 well plates and ³H-thymidineincorporation may be measured using a scintillation counter.

Selection and Screening for Antibodies that Immunospecifically Bind toMembrane-Bound B Lymphocyte Stimulator

Antibodies of the invention (including scFvs and other moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) may be screened in a variety of assays to identifythose antibodies that immunospecifically bind to the membrane-bound formof B Lymphocyte Stimulator. In one particular assay, antibodies thatbind to B Lymphocyte Stimulator on U937 membranes or immobilizedhistidine-tagged B Lymphocyte Stimulator are captured. Other cell linesthat express B Lymphocyte Stimulator that might be useful for testingantibody binding to membrane-bound form of B Lymphocyte Stimulatorinclude, K-562, HL-60 and THP-1 cells. In another assay, antibodies arescreened using ELISAs for those antibodies (or antibody fragments orvariants) that bind to B Lymphocyte Stimulator on U937 membranes or tohistidine-tagged B Lymphocyte Stimulator. In this assay, antibodies areadded to 96 well plates coated with U937 membranes or histidine-tagged BLymphocyte Stimulator and those antibodies or antibody fragments orvariants that bind to the U937 membranes or histidine-tagged BLymphocyte Stimulator are captured. In another assay, antibodies arescreened using ELISAs for those antibodies (or antibody fragments orvariants thereof) that do not bind to biotinylated B LymphocyteStimulator (soluble B Lymphocyte Stimulator) but bind to membrane-boundB Lymphocyte Stimulator, such as, for example, that on membranes fromU937 cells (see Example 2, infra). In these assays, soluble B LymphocyteStimulator (e.g., biotinylated B Lymphocyte Stimulator) andmembrane-bound B Lymphocyte Stimulator (e.g., on U937 membranes) areincubated in separate samples with the same antibodies (or antibodyfragments or variants) and those antibodies (or antibody fragments orvariants) that do not bind to the soluble B Lymphocyte Stimulator(biotinylated B Lymphocyte Stimulator), but bind the membrane-bound BLymphocyte Stimulator (e.g., on U937 membranes) are captured andidentified. In other assays, antibodies are screened using ELISAs todetermine which of the antibodies (or antibody fragments or variants)that bind to histidine-tagged B Lymphocyte Stimulator or membranes fromU937 cells do not cross-react with APRIL, endokine-alpha, VEGI, TRAIL,TNF-alpha, TNF-beta, Fas-L, LIGHT, and PBS (See Example 4, infra).ELISAs can also be used to determine which of the antibodies (orantibody fragments or variants) that bind to histidine-tagged BLymphocyte Stimulator or membranes from U937 cells bind to B LymphocyteStimulator in the presence of TNF-alpha (see Example 4, infra).Antibodies or fragments or variants thereof that immunospecifically bindto the membrane-bound form of B Lymphocyte Stimulator may also be testedfor their affinity for histidine-tagged B Lymphocyte Stimulator usinghigh-throughput BIAcore analysis (see Example 14, infra).

Additionally, antibodies of the invention may be screened against cellsengineered to express an “uncleavable” form of B Lymphocyte Stimulatorin order to determine their specificity for the membrane-bound form of BLymphocyte Stimulator. Mutations in B Lymphocyte Stimulator which mayachieve this result include, but are not limited to, the mutation ordeletion of amino acid residues Lys-132 and/or Arg-133 of the BLymphocyte Stimulator sequence shown in SEQ ID NO:3228. A typicalmutagenesis might include mutation of one or both of residues Lys-132 orArg-133 to alanine residues. Cells expressing such an “uncleavable” formof B Lymphocyte Stimulator provide a profound reagent to use in assayingthe ability of antibodies to bind the membrane-bound form of BLymphocyte Stimulator.

Selection and Screening for Antibodies that Immunospecifically Bind toSoluble and Membrane-Bound B Lymphocyte Stimulator

Antibodies of the invention (including scFvs and other moleculescomprising, or alternately consisting of, antibody fragments orvariants) may be screened in a variety of assays to identify thoseantibodies or antibody fragments or variants that immunospecificallybind to the soluble form and membrane-bound form of B LymphocyteStimulator. In one particular assay, antibodies that bind to immobilizedB Lymphocyte Stimulator are captured. In another assay, antibodies arescreened using ELISAs for those antibodies (or antibody fragments orvariants) that inhibit the binding of soluble B Lymphocyte Stimulator(e.g. soluble bio-B Lymphocyte Stimulator) to IM-9 cells as describedsupra. In other assays, antibodies are screened using ELISAs for thoseantibodies that bind to membranes from U937 cells. Additionally, furtherELISA assays may be performed using techniques known in the art todetermine which antibodies do not cross-react with APRIL,endokine-alpha, VEGI, TRAIL, TNF-alpha, TNF-beta, Fas-L, LIGHT, and PBS,or those antibodies that bind to B Lymphocyte Stimulator in the presenceof TNF-alpha (see Example 4 infra). Antibodies may be assayed inneutralization assays using techniques described herein or otherwiseknown in the art. Antibodies that immunospecifically bind to the solubleand membrane-bound forms of B Lymphocyte Stimulator may also be testedfor their affinity for B Lymphocyte Stimulator using high-throughputBIAcore analysis.

Antibody Conjugates

The present invention encompasses antibodies (including scFvs and othermolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof), recombinantly fused or chemically conjugated(including both covalent and non-covalent conjugations) to aheterologous polypeptide (or portion thereof, preferably at least 10, atleast 20, at least 30, at least 40, at least 50, at least 60, at least70, at least 80, at least 90 or at least 100 amino acids of thepolypeptide) to generate fusion proteins. The fusion does notnecessarily need to be direct, but may occur through linker sequences.For example, antibodies of the invention may be used to targetheterologous polypeptides to particular cell types (e.g., cells ofmonocytic lineage and B-cells), either in vitro or in vivo, by fusing orconjugating the heterologous polypeptides to antibodies of the inventionthat are specific for particular cell surface antigens (e.g.,membrane-bound B Lymphocyte Stimulator on cells of monocytic lineage) orwhich bind antigens that bind particular cell surface receptors (e.g.,TACI, BCMA, BAFF-R located on B cells). Antibodies fused or conjugatedto heterologous polypeptides may also be used in in vitro immunoassaysand purification methods using methods known in the art. See e.g.,Harbor et al., supra, and PCT publication WO 93/2 1232; EP 439,095;Naramura et al., Immunol Lett. 39:91-99 (1994); U.S. Pat. No. 5,474,981;Gillies et al., PNAS 89:1428-1432 (1992); Fell et al., J. Immunol146:2446-2452 (1991), which are incorporated by reference in theirentireties.

In one embodiment, a fusion protein comprises a polypeptide having anamino acid sequence of any one of the VH domains referred to in Table 1,and a heterologous polypeptide. In another embodiment, a fusion proteincomprises a polypeptide having the amino acid sequence of any one of theVH CDR1s referred to in Table 1, and a heterologous polypeptide. Inanother embodiment, a fusion protein comprises a polypeptide having theamino acid sequence of any one of the VH CDR2s referred to in Table 1,and a heterologous polypeptide. In a preferred embodiment, a fusionprotein comprises a polypeptide having the amino acid sequence of anyone of the VH CDR3s referred to in Table 1 (i.e., SEQ ID NOS:2129-3227),and a heterologous polypeptide.

In another embodiment, a fusion protein comprises a polypeptide havingthe amino acid sequence of any one of the VL domains referred to inTable 1, and a heterologous polypeptide. In another embodiment, a fusionprotein comprises a polypeptide having the amino acid sequence of anyone of the VL CDR1s referred to in Table 1, and a heterologouspolypeptide. In yet another embodiment, a fusion protein comprises apolypeptide having the amino acid sequence of any one of the VL CDR2sreferred to in Table 1, and a heterologous polypeptide. In a preferredembodiment, a fusion protein comprises a polypeptide having the aminoacid sequence of any one of the VL CDR3s referred to in Table 1, and aheterologous polypeptide.

In another embodiment, a fusion protein comprises a polypeptide havingthe amino acid sequence of any one of the VH domains referred to inTable 1, and one or more VL domains referred to in Table 1, and aheterologous polypeptide. In another embodiment, a fusion protein of thepresent invention comprises a polypeptide having the amino acid sequenceof any one of the VH CDRs referred to in Table 1, and any one of the VLCDRs referred to in Table 1, and a heterologous polypeptide.

The present invention further includes compositions comprising, oralternatively consisting of, heterologous polypeptides fused orconjugated to antibody fragments. For example, the heterologouspolypeptides may be fused or conjugated to a Fab fragment, Fd fragment,Fv fragment, F(ab)₂ fragment, or a portion thereof. Methods for fusingor conjugating polypeptides to antibody portions are known in the art.See, e.g., U.S. Pat. Nos. 5,336,603; 5,622,929; 5,359,046; 5,349,053;5,447,851; 5,112,946; EP 307,434; EP 367,166; PCT publications WO96/04388; WO 9 1/06570; Ashkenazi et al., Proc. Natl. Acad. Sci. USA 88:10535-10539 (1991); Zheng et al., J. Immunol 154:5590-5600 (1995); andVil et al., Proc. Natl. Acad. Sci. USA 89:11337-11341 (1992) (saidreferences incorporated by reference in their entireties).

Additional fusion proteins of the invention may be generated through thetechniques of gene-shuffling, motif-shuffling, exon-shuffling, and/orcodon-shuffling (collectively referred to as “DNA shuffling”). DNAshuffling may be employed to modulate the activities of antibodies(including scFvs and other molecules comprising, or alternativelyconsisting of, antibody fragments or variants thereof), such methods canbe used to generate antibodies with altered activity (e.g., antibodieswith higher affinities and lower dissociation rates). See, generally,U.S. Pat. Nos. 5,605,793; 5,811,238; 5,830,721; 5,834,252; and5,837,458, and Patten et al., Curr. Opinion Biotechnol. 8:724-33 (1997);Harayama, Trends Biotechnol. 16(2):76-82 (1998); Hansson, et al., J.Mol. Biol. 287:265-76 (1999); and Lorenzo and Blasco, Biotechniques24(2):308-13 (1998) (each of these patents and publications are herebyincorporated by reference in its entirety). In one embodiment,polynucleotides encoding antibodies of the invention may be altered bybeing subjected to random mutagenesis by error-prone PCR, randomnucleotide insertion or other methods prior to recombination. In anotherembodiment, one or more portions of a polynucleotide encoding anantibody which portions immunospecifically bind to B LymphocyteStimulator may be recombined with one or more components, motifs,sections, parts, domains, fragments, etc. of one or more heterologousmolecules.

Moreover, the antibodies of the present invention (including scFvs andother molecules comprising, or alternatively consisting of, antibodyfragments or variants thereof), can be fused to marker sequences, suchas a polypeptides to facilitate purification. In preferred embodiments,the marker amino acid sequence is a hexa-histidine polypeptide, such asthe tag provided in a pQE vector (QIAGEN, Inc., 9259 Eton Avenue,Chatsworth, Calif., 91311), among others, many of which are commerciallyavailable. As described in Gentz et al., Proc. Natl. Acad. Sci. USA86:821-824 (1989), for instance, hexa-histidine provides for convenientpurification of the fusion protein. Other peptide tags useful forpurification include, but are not limited to, the hemagglutinin “HA”tag, which corresponds to an epitope derived from the influenzahemagglutinin protein (Wilson et al., Cell 37:767 (1984)) and the “flag”tag (DYKDDDDK, (SEQ ID No: 3238) Stratagene, La Jolla, Calif.).

The present invention further encompasses antibodies (including scFvsand other molecules comprising, or alternatively consisting of, antibodyfragments or variants thereof), conjugated to a diagnostic ortherapeutic agent. The antibodies can be used diagnostically to, forexample, monitor or prognose the development or progression of a tumoras part of a clinical testing procedure to, e.g., determine the efficacyof a given treatment regimen. Detection can be facilitated by couplingthe antibody to a detectable substance. Examples of detectablesubstances include, but are not limited to, various enzymes, prostheticgroups, fluorescent materials, luminescent materials, bioluminescentmaterials, radioactive materials, positron emitting metals using variouspositron emission tomographies, and nonradioactive paramagnetic metalions. The detectable substance may be coupled or conjugated eitherdirectly to the antibody or indirectly, through an intermediate (suchas, for example, a linker known in the art) using techniques known inthe art. See, for example, U.S. Pat. No. 4,741,900 for metal ions whichcan be conjugated to antibodies for use as diagnostics according to thepresent invention. Examples of suitable enzymes include, but are notlimited to, horseradish peroxidase, alkaline phosphatase,beta-galactosidase, or acetylcholinesterase; examples of suitableprosthetic group complexes include, but are not limited to,streptavidinlbiotin and avidin/biotin; examples of suitable fluorescentmaterials include, but are not limited to, umbelliferone, fluorescein,fluorescein isothiocyanate, rhodamine, dichlorotriazinylaminefluorescein, dansyl chloride or phycoerythrin; an example of aluminescent material includes, but is not limited to, luminol; examplesof bioluminescent materials include, but are not limited to, luciferase,luciferin, and aequorin; and examples of suitable radioactive materialinclude, but are not limited to, iodine (¹³¹I, ¹²⁵I, ¹²³I, ¹²¹I), carbon(¹⁴C), sulfur (³⁵S), tritium (³H), indium (^(115m)In, ^(113m)In, ¹¹²In,¹¹¹In), and technetium (⁹⁹Tc, ^(99m)Tc), thallium (²⁰¹Ti), gallium(⁶⁸Ga, ⁶⁷Ga), palladium (¹⁰³Pd), molybdenum (⁹⁹Mo), xenon (¹³³Xe),fluorine (¹⁸ _(F)), ¹⁵³Sm, ¹⁷⁷Lu, ¹⁵⁹Gd, ¹⁴⁹Pm, ¹⁴⁰La, ¹⁷⁵Yb, ¹⁶⁶Ho,⁹⁰Y, ⁴⁷Sc, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁴²Pr, ¹⁰⁵Rh, ⁹⁷Ru, ⁶⁸Ge, ⁵⁷Co, ⁶⁵Zn, ⁸⁵Sr,³²P, ¹⁵³Gd, ¹⁶⁹Yb, ⁵¹Cr, ⁵⁴Mn, ⁷⁵Se, ¹¹³Sn, and ¹¹⁷Tin.

Further, an antibody of the invention (including an scFv or othermolecule comprising, or alternatively consisting of, antibody fragmentsor variants thereof), may be conjugated to a therapeutic moiety such asa cytotoxin, e.g., a cytostatic or cytocidal agent, a therapeutic agentor a radioactive metal ion, e.g., alpha-emitters such as, for example,²¹³Bi. In specific embodiments, antibodies of the invention are attachedto macrocyclic chelators useful for conjugating radiometal ions,including but not limited to, ¹¹¹In, ¹⁷⁷Lu, ⁹⁰Y, ¹⁶⁶Ho, and ¹⁵³Sm, topolypeptides. In preferred embodiments, the radiometal ion associatedwith the macrocyclic chelators attached to antibodies of the inventionis ¹¹¹In. In preferred embodiments, the radiometal ion associated withthe macrocyclic chelators attached to antibodies of the invention is⁹⁰Y. In specific embodiments, the macrocyclic chelator is1,4,7,10-tetraazacyclododecane-N,N′,N″,N′″-tetraacetic acid (DOTA). Inother specific embodiments, the DOTA is attached to the antibody of theinvention via a linker molecule. Examples of linker molecules useful forconjugating DOTA to a polypeptide are commonly known in the art—see, forexample, DeNardo et al., Clin Cancer Res. 4(10):2483-90, 1998; Petersonet al., Bioconjug. Chem. 10(4):553-7, 1999; and Zimmerman et al, Nucl.Med. Biol. 26(8):943-50, 1999 which are hereby incorporated by referencein their entirety.

A cytotoxin or cytotoxic agent includes any agent that is detrimental tocells and includes such molecules as small molecule toxins andenzymatically active toxins of bacterial, fungal, plant, or animalorigin, or fragments thereof. Examples include, but are not limited to,paclitaxol, cytochalasin B, gramicidin D, ethidium bromide, emetine,mitomycin, etoposide (VP-16), tenoposide, vincristine, vinblastine,colchicin, doxorubicin, daunorubicin, dihydroxy anthracin dione,mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone,glucocorticoids, procaine, tetracaine, lidocaine, propranolol, thymidinekinase, endonuclease, RNAse, and puromycin and fragments, variants orhomologs thereof. Therapeutic agents include, but are not limited to,antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6-thioguanine,cytarabine, 5-fluorouracil decarbazine), alkylating agents (e.g.,mechlorethamine, thioepa chlorambucil, melphalan, carmustine (BSNU) andlomustine (CCNU), cyclophosphamide, busulfan, dibromomannitol,streptozotocin, mitomycin C, and cisdichlorodiamine platinum (II) (DDP)cisplatin), anthracyclines (e.g., daunorubicin (formerly daunomycin) anddoxorubicin), antibiotics (e.g., dactinomycin (formerly actinomycin),bleomycin, mithramycin, and anthramycin (AMC)), and anti-mitotic agents(e.g., vincristine and vinblastine), improsulfan, piposulfan, benzodopa,carboquone, meturedopa, uredopa, altretamine, triethylenemelamine,trietylenephosphoramide, triethylenethiophosphaoramidetrimethylolomelamine, chlornaphazine, cholophosphamide, estramustine,ifosfamide, novembichin, phenesterine, prednimustine, trofosfamide,uracil mustard, chlorozotocin, fotemustine, nimustine, ranimustine,aclacinomysins, azaserine, cactinomycin, calichearnicin, carabicin,carminomycin, carzinophilin, chromomycins, detorubicin,6-diazo-5-oxo-L-norleucine, epirubicin, esorubicin, idarubicin,marcellomycin, mycophenolic acid, nogalamycin, olivomycins, peplomycin,potfiromycin, quelamycin, rodorubicin, streptonigrin, tubercidin,ubenimex, zinostatin, zorubicin, denopterin, pteropterin, trimetrexate,fludarabine, thiamiprine, ancitabine, azacitidine, 6-azauridine,carmofur, dideoxyuridine, doxifluridine, enocitabine, floxuridine, 5-FU,calusterone, dromostanolone propionate, epitiostanol, mepitiostane,testolactone, aminoglutethimide, mitotane, trilostane, frolinic acid,aceglatone, aldophosphamide glycoside, aminolevulinic acid, amsacrine,bestrabucil, bisantrene, edatraxate, defofamine, demecolcine,diaziquone, elfornithine, elliptiniurn acetate, etoglucid, galliumnitrate, hydroxyurea, lentinan, lonidamine, mitoguazone, mopidamol,nitracrine, pentostatin, phenamet, pirarubicin, podophyllinic acid,2-ethylhydrazide, procarbazine, PSKO, razoxane, sizofiran,spirogermanium, tenuazonic acid, triaziquone,2,2′,2″-trichlorotriethylamine, urethan, vindesine, dacarbazine,mannomustine, mitobronitol, mitolactol, pipobroman, gacytosine,arabinoside (“Ara-C”), taxoids, e.g. paclitaxel (TAXOL”, Bristol-MyersSquibb Oncology, Princeton, N.J.) doxetaxel (TAXOTERE”, Rh6ne-PoulencRorer, Antony, France), gemcitabine, ifosfamide, vinorelbine, navelbine,novantrone, teniposide, aminopterin, xeloda, ibandronate, CPT-I 1,topoisomerase inhibitor RFS 2000, difluoromethylornithine (DMFO),retinoic acid, esperamicins, capecitabine, and pharmaceuticallyacceptable salts, acids or derivatives of any of the above. Alsoincluded in this definition are anti-hormonal agents that act toregulate or inhibit hormone action on tumors such as anti-estrogensincluding for example tamoxifen, raloxifene, aromatase inhibiting4(5)-imidazoles, 4 hydroxytamoxifen, trioxifene, keoxifene, LY 117018,onapristone, toremifene (Fareston), and anti-androgens such asflutamide, nilutamide, bicalutamide, leuprolide, and goserelin, andpharmaceutically acceptable salts, acids or derivatives of any of theabove.

Techniques known in the art may be applied to label antibodies of theinvention. Such techniques include, but are not limited to, the use ofbifunctional conjugating agents (see e.g., U.S. Pat. Nos. 5,756,065;5,714,631; 5,696,239; 5,652,361; 5,505,931; 5,489,425; 5,435,990;5,428,139; 5,342,604; 5,274,119; 4,994,560; and 5,808,003; the contentsof each of which are hereby incorporated by reference in its entirety)and direct coupling reactions (e.g., Bolton-Hunter and Chloramine-Treaction).

The antibodies of the invention which are conjugates can be used formodifying a given biological response, the therapeutic agent or drugmoiety is not to be construed as limited to classical chemicaltherapeutic agents. For example, the drug moiety may be a protein orpolypeptide possessing a desired biological activity. Such proteins mayinclude, but are not limited to, for example, a toxin such as abrin,ricin A, alpha toxin, pseudomonas exotoxin, or diphtheria toxin,saporin, momordin, gelonin, pokeweed antiviral protein, alpha-sarcin andcholera toxin; a protein such as tumor necrosis factor,alpha-interferon, beta-interferon, nerve growth factor, platelet derivedgrowth factor, tissue plasminogen activator, an apoptotic agent, e.g.,TNF-alpha, TNF-beta, AIM I (see, International Publication No. WO97/33899), AIM II (see, International Publication No. WO 97/34911), FasLigand (Takahashi et al., Int. Immunol., 6:1567-1574 (1994)), VEGI (see,International Publication No. WO 99/23105), a thrombotic agent or ananti-angiogenic agent, e.g., angiostatin or endostatin; or, biologicalresponse modifiers such as, for example, lymphokines, interleukin-1(IL-1), interleukin-2 (IL-2), interleukin-6 (IL-6), granulocytemacrophage colony stimulating factor (GM-CSF), granulocyte colonystimulating factor (G-CSF), or other growth factors.

In specific embodiments, antibodies of the invention are conjugated orfused to a polypeptide cytotoxin. An example of a suitable polypeptidecytotoxin is a ribosome-inactivating protein. Type Iribosome-inactivating proteins are single-chain proteins, while type IIribosome-inactivating proteins consist of two nonidentical subunits (Aand B chains) joined by a disulfide bond (for a review, see Soria etal., Targeted Diagn. Ther. 7:193 (1992)). In one embodiment, type Iribosome-inactivating proteins that may be used include, but are notlimited to, polypeptides from Saponaria officinalis (e.g., saporin-1,saporin-2, saporin-3, saporin-6), Momordica charantia (e.g, momordin),Byronia dioica (e.g., bryodin, bryodin-2), Trichosanthes kirilowii(e.g., trichosanthin, trichokirin), Gelonium multiflorum (e.g.,gelonin), Phytolacca americana (e.g., pokeweed antiviral protein,pokeweed antiviral protein-II, pokeweed antiviral protein-S), Phytolaccadodecandra (e.g., dodecandrin, Mirabilis antiviral protein).Ribosome-inactivating proteins are described, for example, by Walsh etal., U.S. Pat. No. 5,635,384. In specific embodiments, an antibody ofthe invention is conjugated or fused to a fragment or variant of theribosome inactivating proteins described above, particularly when saidfragment or variant retains activity.

In another embodiment, type II ribosome-inactivating proteins that maybe used according to the invention include, but are not limited to,polypeptides from Ricinus communis (e.g., ricin), Abrus precatorius(e.g., abrin), Adenia digitata (e.g., modeccin). Since type IIribosome-inactivating proteins include a B chain that binds galactosidesand a toxic A chain that depurinates adenosine. Type IIribosome-inactivating protein conjugates should include the A chain.Additional ribosome-inactivating proteins that may be used according tothe invention include, but are not limited to, bouganin, clavin, maizeribosome-inactivating proteins, Vaccaria pyramidataribosome-inactivating proteins, nigrine b, basic nigrine 1, ebuline,racemosine b, luffin-a, luffin-b, luffin-S, and otherribosome-inactivating proteins known to those of skill in the art. See,for example, Bolognesi and Stirpe, International Publication No.WO98/55623, Colnaghi et al., International Publication No. WO97/49726,Hey et al., U.S. Pat. No. 5,635,384, Bolognesi and Stirpe, InternationalPublication No. WO95/07297, Arias et al., International Publication No.WO94/20540, Watanabe et al., J. Biochem. 106:6 977 (1989); Islam et al.,Agric. Biol. Chem. 55:229 (1991), and Gao et al., FEBS Lett. 347:257(1994). In specific embodiments, an antibody of the invention isconjugated or fused to a fragment or variant of the ribosomeinactivating proteins described above, particularly when said fragmentor variant retains activity.

Additional ribosome inactivating proteins (RIPs) that may be conjugatedor fused to antibodies of the invention include, but are not limited to,Type I Plant RIPs such as Pokeweed antiviral proteins, Tritin, Gelonin,Momordin, Saporin, Dianthin, and Maize RIP; Type II Plant RIPs such asRicin, Abrin, Modecin, Viscumin, Volkensin, Cinnamomin, Mistletoe lectinI and Luffangulin (6 kda); Bacterial RIPS such as Shiga toxin, andShiga-like toxin; and Fungal RIPS such as alpha-sarcin, mitogillin, andrestrictocin. In specific embodiments, an antibody of the invention isconjugated or fused to a fragment or variant of the ribosomeinactivating proteins described above, particularly when said fragmentor variant retains activity.

Ricin A homologues that may be conjugated or fused to antibodies of theinvention include, but are not limited to, polypeptides that have thesame amino acid sequence as a protein selected from the group consistingof: type 2 ribosome-inactivating protein cinnamomin III precursor,abrin-d precursor from Indian licorice or fragment thereof, RIPprecursor of Sambucus nigra, RIP bryodin II precursor,alpha-trichosanthin, ribosome-inactivating protein precursor fromSambucus ebulus, karasurin-B, Trichobakin, Beta-Luffin, Beta-galactosidespecific lectin I A chain (MLA; ML-I A), lectin chain A isoform 2 fromViscum album subsp. coloratum, curcin precursor from Jatropha curcas,trichoanguina from snake gourd, ribosome-inactivating protein IRAb fromIris hollandica, Momordin, trichoanguin from Trichosanthes cucumerina,momordin II from Momordica balsamina, gelonin from Gelonium multiflorum,ribosome inactivating protein RIPm from Polygonatum multiflorum,ribosome inactivating protein Euserratin 2 precursor from Euphorbiaserrata, alpha-PAP (pokeweed antiviral protein) from Phytolaccaamericana, ribosome-inactivating protein gynostemmin II from Gynostemmapentaphyllum, trichosanthin, ribosome-inactivating protein 2 fromPhytolacca insularis, bouganin from Bougainvillea spectabilis, antiviralprotein from Clerodendrum aculeatum, anti-viral protein PAP fromPhytolacca acinosa, type I RIP moschatin I, antiviral ribosomeinactivating protein from Chenopodium album, Betavulgin from Betavulgaris subsp. vulgaris, ribosome-inactivating protein benincasin fromBenincasa hispida, ribosome-inactivating protein ME1 from Mirabilisexpansa, MAP—MIRABILIS—Four-o'clock (Marvel-of-peru), RIP amaranthin,amarandin-S from Amaranthus tricolor, ribosome-inactivating protein fromSpinacia oleracea, type 1 RIP musarmin 3 from Muscari, saporin fromSaponaria officinalis, dianthin 30 from Dianthus caryophyllus, andtritin from Triticum aestivum, or biologically active fragments orvariants thereof.

Other toxins that may be conjugated or fused to antibodies of theinvention include, but are not limited to diphtheria toxin, Pseudomonasexotoxin A, Botulinum neurotoxin, Clostridium perfringens epsilon toxin,Conotoxins, and Staphylococcal enterotoxins.

Antibodies of the invention (including scFvs and other moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof), may also be attached to solid supports, which areparticularly useful for immunoassays or purification of the targetantigen. Such solid supports include, but are not limited to, glass,cellulose, polyacrylamide, nylon, polystyrene, polyvinyl chloride orpolypropylene.

Techniques for conjugating a therapeutic moiety to antibodies are wellknown, see, e.g., Arnon et al., “Monoclonal Antibodies ForImmunotargeting Of Drugs In Cancer Therapy”, in Monoclonal AntibodiesAnd Cancer Therapy, Reisfeld et al. (eds.), pp. 243-56 (Alan R. Liss,Inc. 1985); Hellstrom et al., “Antibodies For Drug Delivery”, inControlled Drug Delivery (2nd Ed.), Robinson et al. (eds.), pp. 623-53(Marcel Dekker, Inc. 1987); Thorpe, “Antibody Carriers Of CytotoxicAgents In Cancer Therapy: A Review”, in Monoclonal Antibodies '84:Biological And Clinical Applications, Pinchera et al. (eds.), pp.475-506 (1985); “Analysis, Results, And Future Prospective Of TheTherapeutic Use Of Radiolabeled Antibody In Cancer Therapy”, inMonoclonal Antibodies For Cancer Detection And Therapy, Baldwin et al.(eds.), pp. 303-16 (Academic Press 1985), and Thorpe et al., “ThePreparation And Cytotoxic Properties Of Antibody-Toxin Conjugates”,Immunol. Rev. 62:119-58 (1982).

Alternatively, an antibody of the invention can be conjugated to asecond antibody to form an antibody heteroconjugate as described bySegal in U.S. Pat. No. 4,676,980, which is incorporated herein byreference in its entirety.

An antibody of the invention (including an scFv or and other moleculecomprising, or alternatively consisting of, an antibody fragment orvariant thereof), with or without a therapeutic moiety conjugated to it,administered alone or in combination with cytotoxic factor(s) and/orcytokine(s) can be used as a therapeutic.

Certain forms of antibodies, particularly, IgG3 and IgM antibodies, aswell as IgG1 and IgG2 antibodies to a lesser extent, are able to mediatecellular lysis via the complement cascade. Thus antibodies of theinvention may be used to kill cells expressing Neutrokine-alpha on theirsurface or cells bearing Neutrokine-alpha receptors viacomplement-mediated killing.

Antibodies of the invention may be used to mediate antibody dependentcell-mediated cytotocicity (ADCC) by, for example, Natural Killer (NK)cells. ADCC occurs when antibodies of the IgG1 or IgG3 isotype interactwith FcγRIII receptor on Natural Killer cells triggering an cytotoxicattack by the NK cell on the antibody coated cells. Thus antibodies ofthe invention may be used to kill cells expressing Neutrokine-alpha ontheir surface or cells bearing Neutrokine-alpha receptors via ADCC.

Use of Antibodies for Epitope Mapping

The present invention provides antibodies (including scFvs and othermolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof), that can be used to identify epitopes of BLymphocyte Stimulator. In particular, the antibodies of the presentinvention can be used to identify epitopes of human B LymphocyteStimulator (SEQ ID NOS:3228 and/or 3229) or B Lymphocyte Stimulatorexpressed on human monocytes; murine B Lymphocyte Stimulator (SEQ IDNOS:3230 and/or 3231) or B Lymphocyte Stimulator expressed on murinemonocytes; rat B Lymphocyte Stimulator (either the soluble forms asgiven in SEQ ID NOS:3232, 3233, 3234 and/or 3235 or in a membraneassociated form, e.g., on the surface of rat monocytes); or monkey BLymphocyte Stimulator (e.g., the monkey B Lymphocyte Stimulatorpolypeptides of SEQ ID NOS:3236 and/or 3237, the soluble form of monkeyB Lymphocyte Stimulator, or B Lymphocyte Stimulator expressed on monkeymonocytes)using techniques described herein or otherwise known in theart. Fragments which function as epitopes may be produced by anyconventional means. (See, e.g., Houghten, Proc. Natl. Acad. Sci. USA82:5131-5135 (1985), further described in U.S. Pat. No. 4,631,211.)

Diagnostic Uses of Antibodies

Labeled antibodies of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof)which specifically bind to B Lymphocyte Stimulator can be used fordiagnostic purposes to detect, diagnose, prognose, or monitor diseasesand/or disorders associated with the aberrant expression and/or activityof B Lymphocyte Stimulator or B Lymphocyte Stimulator receptor. Theinvention provides for the detection of aberrant expression of BLymphocyte Stimulator comprising: (a) assaying the expression of BLymphocyte Stimulator in a biological sample from an individual usingone or more antibodies of the invention that immunospecifically binds toB Lymphocyte Stimulator; and (b) comparing the level of B LymphocyteStimulator with a standard level of B Lymphocyte Stimulator, e.g., innormal biological samples, whereby an increase or decrease in theassayed level of B Lymphocyte Stimulator compared to the standard levelof B Lymphocyte Stimulator is indicative of aberrant expression.

By “biological sample” is intended any fluids and/or cells obtained froman individual, body fluid, body tissue, body cell, cell line, tissueculture, or other source which may contain B Lymphocyte Stimulatorprotein or mRNA. Body fluids include, but are not limited to, sera,plasma, urine, synovial fluid, spinal fluid, saliva, and mucous. Tissuessamples may be taken from virtually any tissue in the body. Tissuesamples may also be obtained from autopsy material. Methods forobtaining tissue biopsies and body fluids from mammals are well known inthe art. Where the biological sample is to include mRNA, a tissue biopsyis the preferred source.

The invention also provides for the detection of aberrant expression ofB Lymphocyte Stimulator receptor comprising (a) assaying the expressionof B Lymphocyte Stimulator receptor in a biological sample from anindividual using one or more antibodies or fragments or variants thereofthat immunospecifically binds only to soluble B Lymphocyte Stimulator,but does not inhibit B Lymphocyte Stimulator/B Lymphocyte Stimulatorreceptor binding. Such an antibody, by way of an example that is not tobe construed as limiting, would be one that is able to capture abiotinylated B Lymphocyte Stimulator from solution (see Example 8), butthat would not prevent B Lymphocyte Stimulator from binding to IM-9cells (see Example 3). and (b) comparing the level of B LymphocyteStimulator receptor with a standard level of B Lymphocyte Stimulatorreceptor, e.g., in normal tissue or cell samples, whereby an increase ordecrease in the assayed level of B Lymphocyte Stimulator receptorcompared to the standard level of B Lymphocyte Stimulator receptor isindicative of aberrant expression.

Antibodies of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof)which specifically bind to B Lymphocyte Stimulator can be used fordiagnostic purposes to detect, diagnose, prognose, or monitor autoimmunedisorders and/or immunodeficiencies, and/or diseases or conditionsassociated therewith. The invention provides for the detection ofaberrant expression of B Lymphocyte Stimulator comprising: (a) assayingthe expression of B Lymphocyte Stimulator in a biological sample from anindividual using one or more antibodies of the invention thatimmunospecifically binds to B Lymphocyte Stimulator; and (b) comparingthe level of B Lymphocyte Stimulator with a standard level of BLymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease or decrease in the assayed level of B Lymphocyte Stimulatorcompared to the standard level of B Lymphocyte Stimulator is indicativeof an autoimmune disorder or disease and/or an immunodeficiency. Inspecific embodiments, an increase in the assayed level of B LymphocyteStimulator is indicative of an autoimmune disorder or disease. In otherspecific embodiments, a decrease in the assayed level of B LymphocyteStimulator is indicative of an immunodeficiency.

Antibodies of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof)which specifically bind to B Lymphocyte Stimulator but, do not inhibit BLymphocyte Stimulator/B Lymphocyte Stimulator receptor binding can beused for diagnostic purposes to detect, diagnose, prognose, or monitorautoimmune disorders and/or immunodeficiencies, and/or diseases orconditions associated therewith. The invention provides for thedetection of aberrant expression of B Lymphocyte Stimulator receptorcomprising: (a) assaying the expression of B Lymphocyte Stimulatorreceptor in a biological sample from an individual using one or moreantibodies of the invention that immunospecifically binds to BLymphocyte Stimulator; and (b) comparing the level of B LymphocyteStimulator receptor with a standard level of B Lymphocyte Stimulatorreceptor, e.g., in normal biological samples, whereby an increase ordecrease in the assayed level of B Lymphocyte Stimulator receptorcompared to the standard level of B Lymphocyte Stimulator receptor isindicative of an autoimmune disorder or disease and/or animmunodeficiency. In specific embodiments, an increase in the assayedlevel of B Lymphocyte Stimulator receptor is indicative of an autoimmunedisorder or disease. In other specific embodiments, a decrease in theassayed level of B Lymphocyte Stimulator receptor is indicative of animmunodeficiency.

Autoimmune disorders, diseases, or conditions that may be detected,diagnosed, prognosed, or monitored using the antibodies of the inventioninclude, but are not limited to, autoimmune hemolytic anemia, autoimmuneneonatal thrombocytopenia, idiopathic thrombocytopenia purpura,autoimmune neutropenia, autoimmunocytopenia, hemolytic anemia,antiphospholipid syndrome, dermatitis, gluten-sensitive enteropathy,allergic encephalomyelitis, myocarditis, relapsing polychondritis,rheumatic heart disease, glomerulonephritis (e.g., IgA nephropathy),Multiple Sclerosis, Neuritis, Uveitis Ophthalmia, Polyendocrinopathies,Purpura (e.g., Henloch-Scoenlein purpura), Reiter's Disease, Stiff-ManSyndrome, Autoimmune Pulmonary Inflammation, myocarditis, dilatedcardiomyopathy, IgA glomerulonephritis, dense deposit disease, rheumaticheart disease, Guillain-Barre Syndrome, diabetes mellitus (e.g. Type Idiabetes mellitus or insulin dependent diabetes mellitis), juvenileonset diabetes, and autoimmune inflammatory eye, autoimmune thyroiditis,hypothyroidism (i.e., Hashimoto's thyroiditis, systemic lupuserhythematosus, discoid lupus, Goodpasture's syndrome, Pemphigus,Receptor autoimmunities such as, for example, (a) Graves' Disease, (b)Myasthenia Gravis, and (c) insulin resistance, autoimmune hemolyticanemia, autoimmune thrombocytopenic purpura, rheumatoid arthritis,schleroderma with anti-collagen antibodies, mixed connective tissuedisease, polymyositis/dermatomyositis, pernicious anemia (Addison'sdisease), idiopathic Addison's disease, infertility, glomerulonephritissuch as primary glomerulonephritis and IgA nephropathy, bullouspemphigoid, Sjögren's syndrome, diabetes millitus, and adrenergic drugresistance (including adrenergic drug resistance with asthma or cysticfibrosis), chronic active hepatitis, primary biliary cirrhosis, otherendocrine gland failure, vitiligo, vasculitis, post-MI, cardiotomysyndrome, urticaria, atopic dermatitis, asthma, inflammatory myopathies,and other inflammatory, granulomatous, degenerative, and atrophicdisorders and other disorders such as inflammatory skin diseasesincluding psoriasis and sclerosis, responses associated withinflammatory bowel disease (such as Crohn's disease and ulcerativecolitis), respiratory distress syndrome (including adult respiratorydistress syndrome, ARDS), meningitis, encephalitis, colitis, allergicconditions such as eczema and other conditions involving infiltration ofT cells and chronic inflammatory responses, atherosclerosis, leukocyteadhesion deficiency, Reynaud's syndrome, and immune responses associatedwith acute and delayed hypersensitivity mediated by cytokines andT-lymphocytes typically found in tuberculosis, sarcoidosis,granulomatosis and diseases involving leukocyte diapedesis, centralnervous system (CNS) inflammatory disorder, multiple organ injurysyndrome, antigen-antibody complex mediated diseases, anti-glomerularbasement membrane disease, Lambert-Eaton myasthenic syndrome, Beheetdisease, giant cell arteritis, immune complex nephritis, IgAnephropathy, IgM polyneuropathies or autoimmune thrombocytopenia etc.

In specific embodiments, the present invention encompasses methods andcompositions for detecting, diagnosing and/or prognosing diseases ordisorders associated with hypergammaglobulinemia (e.g., AIDS, autoimmunediseases, and some immunodeficiencies). In other specific embodiments,the present invention encompasses methods and compositions fordetecting, diagnosing and/or prognosing diseases or disorders associatedwith hypogammaglobulinemia (e.g., an immunodeficiency).

Immunodeficiencies that may be detected, diagnosed, prognosed, ormonitored using the antibodies of the invention include, but are notlimited to, severe combined immunodeficiency (SCID)-X linked,SCID-autosomal, adenosine deaminase deficiency (ADA deficiency),X-linked agammaglobulinemia (XLA), Bruton's disease, congenitalagammaglobulinemia, X-linked infantile agammaglobulinemia, acquiredagammaglobulinemia, adult onset agammaglobulinemia, late-onsetagammaglobulinemia, dysgammaglobulinemia, hypogammaglobulinemia,transient hypogammaglobulinemia of infancy, unspecifiedhypogammaglobulinemia, agammaglobulinemia, common variableimmunodeficiency (CVID) (acquired), Wiskott-Aldrich Syndrome (WAS),X-linked immunodeficiency with hyper IgM, non X-linked immunodeficiencywith hyper IgM, selective IgA deficiency, IgG subclass deficiency (withor without IgA deficiency), antibody deficiency with normal or elevatedIgs, immunodeficiency with thymoma, Ig heavy chain deletions, kappachain deficiency, B cell lymphoproliferative disorder (BLPD), selectiveIgM immunodeficiency, recessive agammaglobulinemia (Swiss type),reticular dysgenesis, neonatal neutropenia, severe congenitalleukopenia, thymic alymphoplasia-aplasia or dysplasia withimmunodeficiency, ataxia-telangiectasia, short limbed dwarfism, X-linkedlymphoproliferative syndrome (XLP), Nezelof syndrome-combinedimmunodeficiency with Igs, purine nucleoside phosphorylase deficiency(PNP), MHC Class II deficiency (Bare Lymphocyte Syndrome) and severecombined immunodeficiency.

Elevated levels of soluble B Lymphocyte Stimulator have been observed inthe serum of patients with Systemic Lupus Erythematosus (SLE). Incomparing the sera of 150 SLE patients with that of 38 controlindividuals, it was found that most of the SLE patients had more than 5ng/ml of serum B Lymphocyte Stimulator, more than 30% of SLE patientshad levels greater than 10 ng/ml, and approximately 10% of SLE patientshad serum B Lymphocyte Stimulator levels greater than 20 ng/ml. Incontrast, the majority of normal controls had B Lymphocyte Stimulatorlevels less than 5 ng/ml, and less than 10% had levels higher than 10ng/ml. The elevated levels of B Lymphocyte Stimulator protein in sera ispresent in the soluble form and has biologic activity as assayed by theability to stimulate anti-IgM treated B cells in vitro. SLE patientswith more than 15 ng/ml serum B Lymphocyte Stimulator were also found tohave elevated levels of anti-dsDNA antibodies compared to both normalcontrols and SLE patients with less than 5 ng/ml of serum B LymphocyteStimulator.(unpublished data).

In addition the serum of two subgroups of patients which were positivefor anti-nuclear antibodies (ANA+) but did not meet the formalrequirements of the American College of Rheumatology (ACR) forclassification of SLE were analyzed for B Lymphocyte Stimulator levels.The first subgroup of sera was ANA+ sera that came from patients who didnot present with the clinical impression of SLE. This group had onlyslightly elevated levels of B Lymphocyte Stimulator (˜9 ng/ml BLymphocyte Stimulator). The second subgroup however, which was ANA+ serafrom patients who presented with the clinical impression of SLE, hadsignificantly increased B Lymphocyte Stimulator levels (˜15 ng/ml).These results suggest that an elevated level of B Lymphocyte Stimulatorprecedes the formal fulfillment of the ACR criteria. The ACR criteriaare described in Tan, E. M., et al, Arthritis and Rheumatism25:1271-1277 (1982).

Thus in specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor Systemic LupusErythematosus or conditions associated therewith. The invention providesfor the detection of aberrant expression of B Lymphocyte Stimulatorcomprising: (a) assaying the expression of B Lymphocyte Stimulator in abiological sample of an individual using one or more antibodies of theinvention that immunospecifically binds to B Lymphocyte Stimulator; and(b) comparing the level of B Lymphocyte Stimulator with a standard levelof B Lymphocyte Stimulator, e.g., in normal biological samples, wherebyan increase in the assayed level of B Lymphocyte Stimulator compared tothe standard level of B Lymphocyte Stimulator is indicative of SLE.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor IgA nephropathy orconditions associated therewith. The invention provides for thedetection of aberrant expression of B Lymphocyte Stimulator comprising:(a) assaying the expression of B Lymphocyte Stimulator in a biologicalsample of an individual using one or more antibodies of the inventionthat immunospecifically binds to B Lymphocyte Stimulator; and (b)comparing the level of B Lymphocyte Stimulator with a standard level ofB Lymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease in the assayed level of B Lymphocyte Stimulator compared to thestandard level of B Lymphocyte Stimulator is indicative of IgAnephropathy.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor Sjögren's Syndrome orconditions associated therewith. The invention provides for thedetection of aberrant expression of B Lymphocyte Stimulator comprising:(a) assaying the expression of B Lymphocyte Stimulator in a biologicalsample of an individual using one or more antibodies of the inventionthat immunospecifically binds to B Lymphocyte Stimulator; and (b)comparing the level of B Lymphocyte Stimulator with a standard level ofB Lymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease in the assayed level of B Lymphocyte Stimulator compared to thestandard level of B Lymphocyte Stimulator is indicative of Sjögren'sSyndrome.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor HIV infection orconditions associated therewith (e.g. AIDS). The invention provides forthe detection of aberrant expression of B Lymphocyte Stimulatorcomprising: (a) assaying the expression of B Lymphocyte Stimulator in abiological sample of an individual using one or more antibodies of theinvention that immunospecifically binds to B Lymphocyte Stimulator; and(b) comparing the level of B Lymphocyte Stimulator with a standard levelof B Lymphocyte Stimulator, e.g., in normal biological samples, wherebyan increase in the assayed level of B Lymphocyte Stimulator compared tothe standard level of B Lymphocyte Stimulator is indicative of HIVinfection.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor Myasthenia Gravis orconditions associated therewith. The invention provides for thedetection of aberrant expression of B Lymphocyte Stimulator comprising:(a) assaying the expression of B Lymphocyte Stimulator in a biologicalsample of an individual using one or more antibodies of the inventionthat immunospecifically binds to B Lymphocyte Stimulator; and (b)comparing the level of B Lymphocyte Stimulator with a standard level ofB Lymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease in the assayed level of B Lymphocyte Stimulator compared to thestandard level of B Lymphocyte Stimulator is indicative of MyastheniaGravis.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor idiopathicthrombocytopenic purpura (ITP) or conditions associated therewith. Theinvention provides for the detection of aberrant expression of BLymphocyte Stimulator comprising: (a) assaying the expression of BLymphocyte Stimulator in a biological sample of an individual using oneor more antibodies of the invention that immunospecifically binds to BLymphocyte Stimulator; and (b) comparing the level of B LymphocyteStimulator with a standard level of B Lymphocyte Stimulator, e.g., innormal biological samples, whereby an increase in the assayed level of BLymphocyte Stimulator compared to the standard level of B LymphocyteStimulator is indicative of idiopathic thrombocytopenic purpura (ITP).

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor hemolytic anemia orconditions associated therewith. The invention provides for thedetection of aberrant expression of B Lymphocyte Stimulator comprising:(a) assaying the expression of B Lymphocyte Stimulator in a biologicalsample of an individual using one or more antibodies of the inventionthat immunospecifically binds to B Lymphocyte Stimulator; and (b)comparing the level of B Lymphocyte Stimulator with a standard level ofB Lymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease in the assayed level of B Lymphocyte Stimulator compared to thestandard level of B Lymphocyte Stimulator is indicative of hemolyticanemia.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor thyroiditis orconditions associated therewith. The invention provides for thedetection of aberrant expression of B Lymphocyte Stimulator comprising:(a) assaying the expression of B Lymphocyte Stimulator in a biologicalsample of an individual using one or more antibodies of the inventionthat immunospecifically binds to B Lymphocyte Stimulator; and (b)comparing the level of B Lymphocyte Stimulator with a standard level ofB Lymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease in the assayed level of B Lymphocyte Stimulator compared to thestandard level of B Lymphocyte Stimulator is indicative of thyroiditis.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor Goodpasture'ssyndrome or conditions associated therewith. The invention provides forthe detection of aberrant expression of B Lymphocyte Stimulatorcomprising: (a) assaying the expression of B Lymphocyte Stimulator in abiological sample of an individual using one or more antibodies of theinvention that immunospecifically binds to B Lymphocyte Stimulator; and(b) comparing the level of B Lymphocyte Stimulator with a standard levelof B Lymphocyte Stimulator, e.g., in normal biological samples, wherebyan increase in the assayed level of B Lymphocyte Stimulator compared tothe standard level of B Lymphocyte Stimulator is indicative ofGoodpasture's syndrome.

In other specific embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor multiple sclerosis orconditions associated therewith. The invention provides for thedetection of aberrant expression of B Lymphocyte Stimulator comprising:(a) assaying the expression of B Lymphocyte Stimulator in a biologicalsample of an individual using one or more antibodies of the inventionthat immunospecifically binds to B Lymphocyte Stimulator; and (b)comparing the level of B Lymphocyte Stimulator with a standard level ofB Lymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease in the assayed level of B Lymphocyte Stimulator compared to thestandard level of B Lymphocyte Stimulator is indicative of multiplesclerosis.

In additional embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor Rheumatoid Arthritis.The invention provides for the detection of aberrant expression of BLymphocyte Stimulator comprising: (a) assaying the expression of BLymphocyte Stimulator in a biological sample (e.g., serum and synovialfluid) of an individual using one or more antibodies of the inventionthat immunospecifically binds to B Lymphocyte Stimulator; and (b)comparing the level of B Lymphocyte Stimulator with a standard level ofB Lymphocyte Stimulator, e.g., in normal biological samples, whereby anincrease in the assayed level of B Lymphocyte Stimulator compared to thestandard level of B Lymphocyte Stimulator is indicative of Rheumatoidarthritis.

In additional embodiments, antibodies of the invention whichspecifically bind to B Lymphocyte Stimulator can be used for diagnosticpurposes to detect, diagnose, prognose, or monitor an immune-basedrheumatologic disease, (e.g., SLE, rheumatoid arthritis, CREST syndrome(a variant of schleroderma characterized by calcinosis, Raynaud'sphenomenon, esophageal motility disorders, sclerodactyly, andtelangiectasia.), Seronegative spondyloarthropathy (SpA),Polymyositis/dermatomyositis, Microscopic polyangiitis, HepatitisC-associated arthritis, Takayasu's arteritis, and undifferentiatedconnective tissue disorder). The invention provides for the detection ofaberrant expression of B Lymphocyte Stimulator comprising: (a) assayingthe expression of B Lymphocyte Stimulator in a biological sample (e.g.,serum and synovial fluid) of an individual using one or more antibodiesof the invention that immunospecifically binds to B LymphocyteStimulator; and (b) comparing the level of B Lymphocyte Stimulator witha standard level of B Lymphocyte Stimulator, e.g., in normal biologicalsamples, whereby an increase in the assayed level of B LymphocyteStimulator compared to the standard level of B Lymphocyte Stimulator isindicative of monitor an immune-based rheumatologic disease.

It has been observed, that serum B Lymphocyte Stimulator levelsinversely correlate with nephrotic range proteinuria (>3 gm proteinuriain a 24 hour urine collection) using a sample of 71 SLE patients(p=0.019). Proteinuria was determined in 71 SLE patients within onemonth of phlebotomy for serum B Lymphocyte Stimulator determination.Serum B Lymphocyte Stimulator was classified as low, normal, or highbased on the 5^(th) through 95^(th) percentiles for normal controls.Nephrotic-range proteinuria was inversely correlated with serumNeutrokine-alpha levels. Thus, in specific embodiments, serum levels ofB Lymphocyte Stimulator (determined using one or more antibodies of thepresent invention) in individuals diagnosed with an immune basedrheumatologic disease (e.g., SLE, rheumatoid arthritis, CREST syndrome(a variant of schleroderma characterized by calcinosis, Raynaud'sphenomenon, esophageal motility disorders, sclerodactyly, andtelangiectasia.), seronegative spondyloarthropathy (SpA),polymyositis/dermatomyositis, microscopic polyangiitis, hepatitisC-associated arthritis, Takayasu's arteritis, and undifferentiatedconnective tissue disorder) may be used to determine, diagnose,prognose, or monitor the severity of certain aspects or symptoms of thedisease, such as nephrotic-range proteinuria.

In another specific embodiment, antibodies of the invention are used todiagnose, prognose, treat, or prevent conditions associated with CVID,including, but not limited to, conditions associated with acute andrecurring infections (e.g., pneumonia, bronchitis, sinusitis, otitismedia, sepsis, meningitis, septic arthritis, and osteomyelitis), chroniclung disease, autoimmunity, granulomatous disease, lymphoma, cancers(e.g., cancers of the breast, stomach, colon, mouth, prostate, lung,vagina, ovary, skin, and melanin forming cells (i.e. melanoma),inflammatory bowel disease (e.g., Crohn's disease, ulcerative colitis,and ulcerative proctitis), malabsorption, Hodgkin's disease, andWaldenstrom's macroglobulinemia.

The invention provides a diagnostic assay for diagnosing or prognosing adisease or disorder, comprising: (a) assaying for the level of BLymphocyte Stimulator in a biological sample of an individual using oneor more antibodies of the invention that immunospecifically bind to BLymphocyte Stimulator; and (b) comparing the level of B LymphocyteStimulator with a standard B Lymphocyte Stimulator level, e.g., in abiological sample from a patient without the disease or disorder,whereby an increase or decrease in the assayed B Lymphocyte Stimulatorlevel compared to the standard level of B Lymphocyte Stimulator isindicative of a particular disease or disorder. With respect to cancer,the presence of a relatively high amount of B Lymphocyte Stimulator inbiopsied tissue from an individual may indicate a predisposition for thedevelopment of the disease, or may provide a means for detecting thedisease prior to the appearance of actual clinical symptoms. A moredefinitive diagnosis of this type may allow health professionals toemploy preventative measures or aggressive treatment earlier therebypreventing the development or further progression of the cancer.

In specific embodiments, the presence of a relatively high amount ofmembrane-bound B Lymphocyte Stimulator in a biological sample isindicative of monocytic cell related leukemias or lymphomas, such as,for example acute myelogenous leukemia and/or the severity thereof.

In other specific embodiments, the presence of a relatively high amountof B Lymphocyte Stimulator receptor in a biological sample (asdetermined using antibodies of the invention that bind to soluble BLymphocyte Stimulator, but do not inhibit B Lymphocyte Stimulator/BLymphocyte Stimulator receptor binding) is indicative of B cell relatedleukemias or lymphomas (e.g., chronic lymphocytic leukemia, multiplemyeloma, non-Hodgkin's lymphoma, and Hodgkin's disease), and/or theseverity thereof.

In specific embodiments, the invention provides a diagnostic assay fordiagnosing or prognosing Systemic Lupus Erythematosus, comprising: (a)assaying for the level of B Lymphocyte Stimulator in a biological sampleof an individual using one or more antibodies of the invention thatimmunospecifically bind to B Lymphocyte Stimulator; and (b) comparingthe level of B Lymphocyte Stimulator with a standard B LymphocyteStimulator level, e.g., in a biological sample from a patient withoutSystemic Lupus Erythematosus, whereby an increase in the assayed BLymphocyte Stimulator level compared to the standard level of BLymphocyte Stimulator is indicative of Systemic Lupus Erythematosus.

In specific embodiments, the invention provides a diagnostic assay fordiagnosing or prognosing a Rheumatoid Arthritis, comprising: (a)assaying for the level of B Lymphocyte Stimulator in a biological sampleof an individual using one or more antibodies of the invention thatimmunospecifically bind to B Lymphocyte Stimulator; and (b) comparingthe level of B Lymphocyte Stimulator with a standard B LymphocyteStimulator level, e.g., in a biological sample from a patient withoutRheumatoid Arthritis, whereby an increase or decrease in the assayed BLymphocyte Stimulator level compared to the standard level of BLymphocyte Stimulator is indicative of Rheumatoid Arthritis.

The invention provides a diagnostic assay for diagnosing or prognosing adisease or disorder, comprising: (a) assaying for the level of BLymphocyte Stimulator receptor in cells or a tissue sample of anindividual using one or more antibodies of the invention thatimmunospecifically binds only to soluble B Lymphocyte Stimulator, butdoes not neutralize B Lymphocyte Stimulator/B Lymphocyte Stimulatorreceptor binding; and (b) comparing the level of B Lymphocyte Stimulatorreceptor with a standard B Lymphocyte Stimulator receptor level, e.g.,in a tissue sample from a patient without the disease or disorder,whereby an increase or decrease in the assayed B Lymphocyte Stimulatorreceptor level compared to the standard level of B Lymphocyte Stimulatorreceptor is indicative of a particular disease or disorder. With respectto cancer, the presence of a relatively high amount of B LymphocyteStimulator receptor in biopsied tissue from an individual may indicate apredisposition for the development of the disease, or may provide ameans for detecting the disease prior to the appearance of actualclinical symptoms. A more definitive diagnosis of this type may allowhealth professionals to employ preventative measures or aggressivetreatment earlier thereby preventing the development or furtherprogression of the cancer.

Antibodies of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof) canbe used to assay protein levels in a biological sample using classicalimmunohistological methods as described herein or as known to those ofskill in the art (e.g., see Jalkanen, et al., J. Cell. Biol. 101:976-985(1985); Jalkanen, et al., J. Cell. Biol. 105:3087-3096 (1987)). Otherantibody-based methods useful for detecting protein gene expressioninclude immunoassays, such as the enzyme linked immunosorbent assay(ELISA) and the radioimmunoassay (RIA). Suitable antibody assay labelsare known in the art and include enzyme labels, such as, glucoseoxidase, alkaline phosphatase, and horseradish peroxidase;radioisotopes, such as iodine (¹²¹I, ¹²³I, ¹²⁵I, ¹³¹I), carbon (¹⁴C),sulfur (³⁵S), tritium (³H), indium (¹¹¹In, ¹¹²In, ^(113m)In, ^(115m)In),technetium (⁹⁹Tc, ^(99m)Tc), thallium (²⁰¹Ti), gallium (⁶⁸Ga, ⁶⁷Ga),palladium (¹⁰³Pd) molybdenum (⁹⁹Mo), xenon (¹³³Xe), fluorine (¹⁸F),¹⁵³Sm, ¹⁷⁷Lu, ¹⁵⁹Gd, ¹⁴⁹Pm, ¹⁴⁰La, ¹⁷⁵Yb, ¹⁶⁶Ho, ⁹⁰Y, ⁴⁷Sc, ¹⁸⁶Re,¹⁸⁸Re, ¹⁴²Pr, ¹⁰⁵Rh, and ⁹⁷Ru; luminescent labels, such as luminol; andfluorescent labels, such as fluorescein and rhodamine, and biotin.

One aspect of the invention is the detection and diagnosis of a diseaseor disorder associated with aberrant expression of B LymphocyteStimulator or B Lymphocyte Stimulator receptor in an animal, preferablya mammal and most preferably a human. In one embodiment, diagnosiscomprises: a) administering (for example, parenterally, subcutaneously,or intraperitoneally) to a subject an effective amount of a labeledantibody of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof)that immunospecifically binds to B Lymphocyte Stimulator; b) waiting fora time interval following the administering for permitting the labeledantibody to preferentially concentrate at sites in the subject where BLymphocyte Stimulator is expressed (and for unbound labeled molecule tobe cleared to background level); c) determining background level; and d)detecting the labeled antibody in the subject, such that detection oflabeled antibody or fragment thereof above the background level andabove or below the level observed in a person without the disease ordisorder indicates that the subject has a particular disease or disorderassociated with aberrant expression of B Lymphocyte Stimulator or BLymphocyte Stimulator receptor. Background level can be determined byvarious methods including, comparing the amount of labeled moleculedetected to a standard value previously determined for a particularsystem.

It will be understood in the art that the size of the subject and theimaging system used will determine the quantity of imaging moiety neededto produce diagnostic images. In the case of a radioisotope moiety, fora human subject, the quantity of radioactivity injected will normallyrange from about 5 to 20 millicuries of ⁹⁹Tc. The labeled antibody willthen preferentially accumulate at the location of cells which containthe specific protein. In vivo tumor imaging is described in S. W.Burchiel et al., “Immunopharmacokinetics of Radiolabeled Antibodies andTheir Fragments.” (Chapter 13 in Tumor Imaging: The RadiochemicalDetection of Cancer, S. W. Burchiel and B. A. Rhodes, eds., MassonPublishing Inc. (1982).

Depending on several variables, including the type of label used and themode of administration, the time interval following the administrationfor permitting the labeled molecule to preferentially concentrate atsites in the subject and for unbound labeled molecule to be cleared tobackground level is 6 to 48 hours or 6 to 24 hours or 6 to 12 hours. Inanother embodiment the time interval following administration is 5 to 20days or 5 to 10 days.

In an embodiment, monitoring of the disease or disorder is carried outby repeating the method for diagnosing the disease or disorder, forexample, one month after initial diagnosis, six months after initialdiagnosis, one year after initial diagnosis, etc.

Presence of the labeled molecule can be detected in the patient usingmethods known in the art for in vivo scanning These methods depend uponthe type of label used. Skilled artisans will be able to determine theappropriate method for detecting a particular label. Methods and devicesthat may be used in the diagnostic methods of the invention include, butare not limited to, computed tomography (CT), whole body scan such asposition emission tomography (PET), magnetic resonance imaging (MRI),and sonography.

In a specific embodiment, the molecule is labeled with a radioisotopeand is detected in the patient using a radiation responsive surgicalinstrument (Thurston et al., U.S. Pat. No. 5,441,050). In anotherembodiment, the molecule is labeled with a fluorescent compound and isdetected in the patient using a fluorescence responsive scanninginstrument. In another embodiment, the molecule is labeled with apositron emitting metal and is detected in the patient using positronemission-tomography. In yet another embodiment, the molecule is labeledwith a paramagnetic label and is detected in a patient using magneticresonance imaging (MRI).

Immunophenotyping

The antibodies of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof) maybe utilized for immunophenotyping of cell lines and biological samplesby their B Lymphocyte Stimulator expression or B Lymphocyte Stimulatorreceptor expression. Various techniques can be utilized usingantibodies, fragments, or variants of the invention to screen forcellular populations (i.e., immune cells, particularly monocytic cellsor B-cells) expressing B Lymphocyte Stimulator or B LymphocyteStimulator receptor, and include magnetic separation usingantibody-coated magnetic beads, “panning” with antibody attached to asolid matrix (i.e., plate), and flow cytometry (see, e.g., U.S. Pat. No.5,985,660; and Morrison et al., Cell, 96:737-49 (1999)).

These techniques allow for the screening of particular populations ofcells, such as might be found with hematological malignancies (i e.,minimal residual disease (MRD) in acute leukemic patients) and“non-self” cells in transplantations to prevent Graft-versus-HostDisease (GVHD). Alternatively, these techniques allow for the screeningof hematopoietic stem and progenitor cells capable of undergoingproliferation and/or differentiation, as might be found in humanumbilical cord blood.

In one embodiment, antibodies of the invention (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) are used to identify cells of monocytic or B cellorigin.

Use of Antibodies to Identify, Purify and/or Target B LymphocyteStimulator Protein and/or B Lymphocyte Stimulator or B LymphocyteStimulator Receptor Expressing Cells

The antibodies of the invention may be used to identify, purify andtarget B Lymphocyte Stimulator protein and/or B Lymphocyte Stimulator orB Lymphocyte Stimulator receptor expressing cells. For example, theantibodies of the invention when coupled to a detectable label can beused to identify cells that express B Lymphocyte Stimulator. Morespecifically, the antibodies of the invention can be used inimmunohistochemistry or in flow cytometry. Additionally, the antibodiesof the invention may be used to purified B Lymphocyte Stimulatorproteins or portions thereof from cells expressing B LymphocyteStimulator. In more specific embodiments, antibodies of the inventionmay be conjugated (directly or indirectly) to a detectable label such afluorescent label and used in fluorescence activated cell sorting(FACS). Alternatively, antibodies of the invention may be conjugated(directly or indirectly) to a magnetic particle or bead and used inmagnetic cell sorting (MACS).

Antibodies of the invention may be used to target therapeutic or toxicmoieties to B Lymphocyte Stimulator or B Lymphocyte Stimulator Receptorexpressing cells (see Antibody Conjugates section above). In a specificembodiment, antibodies of the invention may also be used to kill BLymphocyte Stimulator or B Lymphocyte Stimulator Receptor expressingcells. In one embodiment, antibodies of the invention are linked to acytotoxic moiety which kills B Lymphocyte Stimulator or B LymphocyteStimulator Receptor expressing cells that have been contacted with suchan antibody conjugate.

Certain forms of antibodies, particularly, IgG3 and IgM antibodies, aswell as IgG1 and IgG2 antibodies to a lesser extent, are able to mediatecellular lysis via the complement cascade. Thus antibodies of theinvention may be used to kill B Lymphocyte Stimulator or B LymphocyteStimulator Receptor expressing cells via complement-mediated killing.

Antibodies of the invention may be used to mediate antibody dependentcell-mediated cytotocicity (ADCC) by, for example, Natural Killer (NK)cells. ADCC occurs when antibodies of the IgG1 or IgG3 isotype interactwith FcγRIII receptor on Natural Killer cells triggering an cytotoxicattack by the NK cell on the antibody coated cells. Thus antibodies ofthe invention may be used to kill B Lymphocyte Stimulator or BLymphocyte Stimulator Receptor expressing cells via ADCC.

Therapeutic Uses of Antibodies

The present invention is further directed to antibody-based therapieswhich involve administering antibodies of the invention (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) to an animal, preferably a mammal, and mostpreferably a human, patient for treating one or more of the discloseddiseases, disorders, or conditions. Therapeutic compounds of theinvention include, but are not limited to, antibodies of the inventionand nucleic acids encoding antibodies (and anti-idiotypic antibodies) ofthe invention as described herein. The antibodies of the invention canbe used to treat, ameliorate or prevent diseases, disorders orconditions associated with aberrant expression and/or activity of BLymphocyte Stimulator or B Lymphocyte Stimulator receptor, including,but not limited to, any one or more of the diseases, disorders, orconditions described herein. The treatment and/or prevention ofdiseases, disorders, or conditions associated with aberrant B LymphocyteStimulator expression and/or activity or aberrant B LymphocyteStimulator receptor expression and/or activity includes, but is notlimited to, alleviating symptoms associated with those diseases,disorders or conditions. Antibodies of the invention may be provided inpharmaceutically acceptable compositions as known in the art or asdescribed herein.

Antibodies of the present invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof)that function as agonists or antagonists of B Lymphocyte Stimulator,preferably of B Lymphocyte Stimulator-induced signal transduction, canbe administered to an animal to treat, prevent or ameliorate a diseaseor disorder associated with aberrant B Lymphocyte Stimulator expression,lack of B Lymphocyte Stimulator function, aberrant B LymphocyteStimulator receptor expression, or lack of B Lymphocyte Stimulatorreceptor function. For example, antibodies of the invention whichdisrupt the interaction between B Lymphocyte Stimulator and its receptormay be administered to an animal to treat, prevent or ameliorate adisease or disorder associated with aberrant B Lymphocyte Stimulatorexpression, excessive B Lymphocyte Stimulator function, aberrant BLymphocyte Stimulator receptor expression, or excessive of B LymphocyteStimulator receptor function. Antibodies of the invention which do notprevent B Lymphocyte Stimulator from binding its receptor but inhibit ordownregulate B Lymphocyte Stimulator-induced signal transduction can beadministered to an animal to treat, prevent or ameliorate a disease ordisorder associated with aberrant B Lymphocyte Stimulator expression,excessive B Lymphocyte Stimulator function, aberrant B LymphocyteStimulator receptor expression, or excessive B Lymphocyte Stimulatorreceptor function. In particular, antibodies of the present inventionwhich prevent B Lymphocyte Stimulator-induced signal transduction byspecifically recognizing the unbound B Lymphocyte Stimulator,receptor-bound B Lymphocyte Stimulator or both unbound andreceptor-bound B Lymphocyte Stimulator can be administered to an animalto treat, prevent or ameliorate a disease or disorder associated withaberrant B Lymphocyte Stimulator expression, excessive B LymphocyteStimulator function, aberrant B Lymphocyte Stimulator receptorexpression, or excessive B Lymphocyte Stimulator receptor function. Theability of an antibody of the invention to inhibit or downregulate BLymphocyte Stimulator-induced signal transduction may be determined bytechniques described herein or otherwise known in the art. For example,B Lymphocyte Stimulator-induced receptor activation and the activationof signaling molecules can be determined by detecting thephosphorylation (e.g., tyrosine or serine/threonine) of the receptor ora signaling molecule by immunoprecipitation followed by western blotanalysis (for example, as described herein).

In a specific embodiment, an antibody of the present invention(including molecules comprising, or alternatively consisting of,antibody fragments or variants thereof) that inhibits or downregulates BLymphocyte Stimulator activity by at least 95%, at least 90%, at least85%, at least 80%, at least 75%, at least 70%, at least 60%, at least50%, at least 45%, at least 40%, at least 45%, at least 35%, at least30%, at least 25%, at least 20%, or at least 10% relative to BLymphocyte Stimulator activity in absence of the antibody isadministered to an animal to treat, prevent or ameliorate a disease ordisorder associated with aberrant B Lymphocyte Stimulator expression,excessive B Lymphocyte Stimulator function, aberrant B LymphocyteStimulator receptor expression, or excessive B Lymphocyte Stimulatorreceptor function. In another embodiment, a combination of antibodies, acombination of antibody fragments, a combination of antibody variants,or a combination of antibodies, antibody fragments, and/or variants thatinhibit or downregulate B Lymphocyte Stimulator activity by at least95%, at least 90%, at least 85%, at least 80%, at least 75%, at least70%, at least 65%, at least 60%, at least 55%, at least 50%, at least45%, at least 40%, at least 45%, at least 35%, at least 30%, at least25%, at least 20%, or at least 10% relative to B Lymphocyte Stimulatoractivity in absence of said antibodies, antibody fragments, and/orantibody variants are administered to an animal to treat, prevent orameliorate a disease or disorder associated with aberrant B LymphocyteStimulator expression, excessive B Lymphocyte Stimulator function,aberrant B Lymphocyte Stimulator receptor expression, or excessive BLymphocyte Stimulator receptor function.

Further, antibodies of the present invention (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) which activate B Lymphocyte Stimulator-induced signaltransduction can be administered to an animal to treat, prevent orameliorate a disease or disorder associated with aberrant B LymphocyteStimulator expression, lack of B Lymphocyte Stimulator function,aberrant B Lymphocyte Stimulator receptor expression, or lack of BLymphocyte Stimulator receptor function. These antibodies may potentiateor activate either all or a subset of the biological activities of BLymphocyte Stimulator-mediated receptor activation, for example, byinducing multimerization of B Lymphocyte Stimulator and/ormultimerization of the receptor. The antibodies of the invention may beadministered with or without being pre-complexed with B LymphocyteStimulator. In a specific embodiment, an antibody of the presentinvention that increases B Lymphocyte Stimulator activity by at least5%, at least 10%, at least 15%, at least 20%, at least 25%, at least30%, at least 35%, at least 40%, at least 45%, at least 50%, at least55%, at least 60%, at least 65%, at least 70%, at least 75%, at least80%, at least 85%, at least 90%, at least 95%, or at least 99% relativeto B Lymphocyte Stimulator activity in absence of the antibody isadministered to an animal to treat, prevent or ameliorate a disease ordisorder associated with aberrant B Lymphocyte Stimulator expression,lack of B Lymphocyte Stimulator function, aberrant B LymphocyteStimulator receptor expression, or lack of B Lymphocyte Stimulatorreceptor function. In another embodiment, a combination of antibodies, acombination of antibody fragments, a combination of antibody variants,or a combination of antibodies, antibody fragments and/or antibodyvariants that increase B Lymphocyte Stimulator activity by at least 5%,at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, atleast 35%, at least 40%, at least 45%, at least 50%, at least 55%, atleast 60%, at least 65%, at least 70%, at least 75%, at least 80%, atleast 85%, at least 90%, at least 95%, or at least 99% relative to BLymphocyte Stimulator activity in absence of the said antibodies orantibody fragments and/or antibody variants is administered to an animalto treat, prevent or ameliorate a disease or disorder associated withaberrant B Lymphocyte Stimulator expression or lack of B LymphocyteStimulator function or aberrant B Lymphocyte Stimulator receptorexpression or lack of B Lymphocyte Stimulator receptor function.

One or more antibodies of the present invention (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to B LymphocyteStimulator may be used locally or systemically in the body as atherapeutic. The antibodies of this invention (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) may also be advantageously utilized in combinationwith other monoclonal or chimeric antibodies, or with lymphokines orhematopoietic growth factors (such as, e.g., IL-2, IL-3 and IL-7), forexample, which serve to increase the number or activity of effectorcells which interact with the antibodies.

The antibodies of the invention (including molecules comprising, oralternatively consisting of, antibody fragments or variants thereof) maybe administered alone or in combination with other types of treatments(e.g., radiation therapy, chemotherapy, hormonal therapy, immunotherapy,anti-tumor agents, anti-angiogenesis and anti-inflammatory agents).Generally, administration of products of a species origin or speciesreactivity (in the case of antibodies) that is the same species as thatof the patient is preferred. Thus, in a preferred embodiment, humanantibodies, fragments, or variants, (e.g., derivatives), or nucleicacids, are administered to a human patient for therapy or prophylaxis.

It is preferred to use high affinity and/or potent in vivo inhibitingand/or neutralizing antibodies of the invention (including moleculescomprising, or alternatively consisting of, antibody fragments orvariants thereof) that immunospecifically bind to B LymphocyteStimulator, or polynucleotides encoding antibodies thatimmunospecifically bind to B Lymphocyte Stimulator, for bothimmunoassays directed to and therapy of disorders related to BLymphocyte Stimulator polynucleotides or polypeptides, includingfragments thereof. Such antibodies will preferably have an affinity forB Lymphocyte Stimulator and/or B Lymphocyte Stimulator fragments.Preferred binding affinities include those with a dissociation constantor K_(D) less than or equal to 5×10⁻² M, 10⁻² M, 5×10⁻³ M, 10⁻³ M,5×10⁻⁴ M, 10⁻⁴ M, 5×10⁻⁵ M, or 10⁻⁵ M. More preferably, antibodies ofthe invention bind B Lymphocyte Stimulator polypeptides or fragments orvariants thereof with a dissociation constant or K_(D) less than orequal to 5×10⁻⁶ M, 10⁻⁶ M, 5×10⁻⁷ M, 10⁻⁷ M, 5×10⁻⁸ M, or 10⁻⁸ M. Evenmore preferably, antibodies of the invention bind B LymphocyteStimulator polypeptides or fragments or variants thereof with adissociation constant or K_(D) less than or equal to 5×10⁻⁹ M, 10⁻⁹ M,5×10⁻¹⁰ M, 10⁻¹⁰M, 5×10⁻¹¹ M, 10⁻¹¹ M, 5×10⁻¹² M, 10⁻¹² M, 5×⁻¹³ M,10⁻¹³M, 5×10⁻¹⁴ M, 10⁻¹⁴ M, 5×10⁻¹⁵ M, or 10⁻¹⁵ M. The invention encompassesantibodies that bind B Lymphocyte Stimulator polypeptides with adissociation constant or K_(D) that is within any one of the ranges thatare between each of the individual recited values.

In a preferred embodiment, antibodies of the invention neutralize BLymphocyte Stimulator activity. In another preferred embodiment,antibodies of the invention inhibit B cell proliferation.

In a preferred embodiment, antibodies of the invention (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) inhibit or reduce binding of the soluble form of BLymphocyte Stimulator to a B Lymphocyte Stimulator receptor. In anotherpreferred embodiment antibodies of the invention inhibit or reduce Bcell proliferation induced by the soluble form of B LymphocyteStimulator. In another preferred embodiment antibodies of the inventioninhibit or reduce immunoglobulin production induced by the soluble formof B Lymphocyte Stimulator.

In a preferred embodiment, antibodies of the invention (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) inhibit or reduce binding of membrane-bound BLymphocyte Stimulator to a B Lymphocyte Stimulator receptor. In anotherpreferred embodiment, antibodies of the invention inhibit or reduce Bcell proliferation induced by the membrane-bound form of B LymphocyteStimulator. In another preferred embodiment, antibodies of the inventioninhibit or reduce immunoglobulin production induced by the membranebound form of B Lymphocyte Stimulator.

In a preferred embodiment, antibodies of the invention (includingmolecules comprising, or alternatively consisting of, antibody fragmentsor variants thereof) inhibit or reduce binding of both the soluble andmembrane-bound forms of B Lymphocyte Stimulator to a B LymphocyteStimulator receptor. In another preferred embodiment, antibodies of theinvention inhibit or reduce B cell proliferation induced by either orboth forms of B Lymphocyte Stimulator. In another preferred embodiment,antibodies of the invention inhibit or reduce immunoglobulin productioninduced by either or both forms of B Lymphocyte Stimulator.

In one embodiment, the invention provides a method of deliveringantibody conjugates of the invention to targeted cells, such as, forexample, monocytic cells expressing the membrane-bound form of BLymphocyte Stimulator, or B cells expressing a B Lymphocyte Stimulatorreceptor.

In one embodiment, the invention provides a method for the specificdelivery of antibodies and antibody conjugates of the invention to cellsby administering molecules of the invention that are associated withheterologous polypeptides or nucleic acids. In one example, theinvention provides a method for delivering a therapeutic protein intothe targeted cell. In another example, the invention provides a methodfor delivering a single stranded nucleic acid (e.g., antisense orribozymes) or double stranded nucleic acid (e.g., DNA that can integrateinto the cell's genome or replicate episomally and that can betranscribed) into the targeted cell.

In another embodiment, the invention provides a method for the specificdestruction of cells (e.g., the destruction of tumor cells) byadministering antibodies or antibody conjugates of the invention (e.g.,antibodies conjugated with radioisotopes, toxins, or cytotoxicprodrugs). In a specific embodiment, the invention provides a method forthe specific destruction of cells of monocytic lineage (e.g., monocyticcell related leukemias or lymphomas, such as, for example acutemyelogenous leukemia) by administering antibodies or antibody conjugatesof the invention (e.g., antibodies conjugated with radioisotopes,toxins, or cytotoxic prodrugs) that immunospecifically bind themembrane-bound form of B Lymphocyte Stimulator. In another specificembodiment, the invention provides a method for the specific destructionof cells of B cell lineage (e.g., B cell related leukemias or lymphomas(e.g., chronic lymphocytic leukemia, multiple myeloma, non-Hodgkin'slymphoma, and Hodgkin's disease) by administering antibodies or antibodyconjugates of the invention (e.g., antibodies conjugated withradioisotopes, toxins, or cytotoxic prodrugs) that bind soluble BLymphocyte Stimulator, but do not inhibit B Lymphocyte Stimulatorbinding to a B Lymphocyte Stimulator receptor on B cells.

In another preferred embodiment antibodies of the invention (includingantibody fragments and variants) promote or enhance B cell proliferationinduced by the soluble form of B Lymphocyte Stimulator. In anotherpreferred embodiment, antibodies of the invention (including antibodyfragments and variants) promote or enhance B cell proliferation inducedby the membrane or soluble form of APRIL. In another preferredembodiment antibodies of the invention (including antibody fragments andvariants) increase or enhance immunoglobulin production induced by thesoluble form of B Lymphocyte Stimulator. In another preferred embodimentantibodies of the invention (including antibody fragments and variants)increase or enhance immunoglobulin production induced by the membranebound or soluble form of APRIL. In another preferred embodimentantibodies of the invention (including antibody fragments and variants)increase or enhance immunoglobulin production in response to T celldependent immunogens. In another preferred embodiment antibodies of theinvention (including antibody fragments and variants, and anti-antibodyantibodies) increase or enhance immunoglobulin production in response toT cell independent immunogens.

In another embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent or ameliorateimmune disorders Immune disorders include, but are not limited to,autoimmune disorders (e.g., arthritis, graft rejection, Hashimoto'sthyroiditis, insulin-dependent diabetes, lupus, idiopathicthrombocytopenic purpura, systemic lupus erythrematosus and multiplesclerosis), elective IgA deficiency, ataxia-telangiectasia, commonvariable immunodeficiency (CVID), X-linked agammaglobulinemia, severecombined immunodeficiency (SCID), Wiskott-Aldrich syndrome, idiopathichyper-eosinophilic syndrome, monocytic leukemoid reaction, monocyticleukocytosis, monocytic leukopenia, monocytopenia, monocytosis, andgraft or transplant rejection.

As discussed herein, antibodies and antibody compositions of theinvention, may be used to treat, prevent, ameliorate, diagnose orprognose various immune system-related disorders and/or conditionsassociated with these disorders, in mammals, preferably humans. Manyautoimmune disorders result from inappropriate recognition of self asforeign material by immune cells. This inappropriate recognition resultsin an immune response leading to the destruction of the host tissue.Therefore, the administration of antibody and antibody compositions ofthe invention that can inhibit an immune response, particularly theproliferation of B cells and/or the production of immunoglobulins, maybe an effective therapy in treating and/or preventing autoimmunedisorders. Thus, in preferred embodiments, antibodies and antibodycompositions of the invention are used to treat, prevent, ameliorate,diagnose and/or prognose an autoimmune disorder, or condition(s)associated with such disorder.

Autoimmune disorders and conditions associated with these disorders thatmay be treated, prevented, ameliorated, diagnosed and/or prognosed withthe therapeutic and pharmaceutical compositions of the inventioninclude, but are not limited to, autoimmune hemolytic anemia, autoimmuneneonatal thrombocytopenia, idiopathic thrombocytopenia purpura,autoimmune neutropenia, autoimmunocytopenia, hemolytic anemia,antiphospholipid syndrome, dermatitis, gluten-sensitive enteropathy,allergic encephalomyelitis, myocarditis, relapsing polychondritis,rheumatic heart disease, glomerulonephritis (e.g., IgA nephropathy),Multiple Sclerosis, Neuritis, Uveitis Ophthalmia, Polyendocrinopathies,Purpura (e.g., Henloch-Scoenlein purpura), Reiter's Disease, Stiff-ManSyndrome, Autoimmune Pulmonary Inflammation, myocarditis, IgAglomerulonephritis, dense deposit disease, rheumatic heart disease,Guillain-Barre Syndrome, insulin dependent diabetes mellitis, andautoimmune inflammatory eye disease.

Additional autoimmune disorders and conditions associated with thesedisorders that may be treated, prevented, ameliorated, diagnosed and/orprognosed with the therapeutic and pharmaceutical compositions of theinvention include, but are not limited to, autoimmune thyroiditis,hypothyroidism (i.e., Hashimoto's thyroiditis) (often characterized,e.g., by cell-mediated and humoral thyroid cytotoxicity), systemic lupuserhythematosus (often characterized, e.g., by circulating and locallygenerated immune complexes), discoid lupus, Goodpasture's syndrome(often characterized, e.g., by anti-basement membrane antibodies),Pemphigus (often characterized, e.g., by epidermal acantholyticantibodies), Receptor autoimmunities such as, for example, (a) Graves'Disease (often characterized, e.g., by TSH receptor antibodies), (b)Myasthenia Gravis (often characterized, e.g., by acetylcholine receptorantibodies), and (c) insulin resistance (often characterized, e.g., byinsulin receptor antibodies), autoimmune hemolytic anemia (oftencharacterized, e.g., by phagocytosis of antibody-sensitized RBCs),autoimmune thrombocytopenic purpura (often characterized, e.g., byphagocytosis of antibody-sensitized platelets.

Additional autoimmune disorders and conditions associated with thesedisorders that may be treated, prevented, ameliorated, diagnosed and/orprognosed with the therapeutic and pharmaceutical compositions of theinvention include, but are not limited to, rheumatoid arthritis (oftencharacterized, e.g., by immune complexes in joints), schleroderma withanti-collagen antibodies (often characterized, e.g., by nucleolar andother nuclear antibodies), mixed connective tissue disease (oftencharacterized, e.g., by antibodies to extractable nuclear antigens(e.g., ribonucleoprotein)), polymyositis/dermatomyositis (oftencharacterized, e.g., by nonhistone ANA), pernicious anemia (oftencharacterized, e.g., by antiparietal cell, microsomes, and intrinsicfactor antibodies), idiopathic Addison's disease (often characterized,e.g., by humoral and cell-mediated adrenal cytotoxicity, infertility(often characterized, e.g., by antispermatozoal antibodies),glomerulonephritis (often characterized, e.g., by glomerular basementmembrane antibodies or immune complexes) such as primaryglomerulonephritis and IgA nephropathy, bullous pemphigoid (oftencharacterized, e.g., by IgG and complement in basement membrane),Sjögren's syndrome (often characterized, e.g., by multiple tissueantibodies, and/or a specific nonhistone ANA (SS-B)), diabetes millitus(often characterized, e.g., by cell-mediated and humoral islet cellantibodies), and adrenergic drug resistance (including adrenergic drugresistance with asthma or cystic fibrosis) (often characterized, e.g.,by beta-adrenergic receptor antibodies), chronic active hepatitis (oftencharacterized, e.g., by smooth muscle antibodies), primary biliarycirrhosis (often characterized, e.g., by mitchondrial antibodies), otherendocrine gland failure (often characterized, e.g., by specific tissueantibodies in some cases), vitiligo (often characterized, e.g., bymelanocyte antibodies), vasculitis (often characterized, e.g., by Ig andcomplement in vessel walls and/or low serum complement), post-MI (oftencharacterized, e.g., by myocardial antibodies), cardiotomy syndrome(often characterized, e.g., by myocardial antibodies), urticaria (oftencharacterized, e.g., by IgG and IgM antibodies to IgE), atopicdermatitis (often characterized, e.g., by IgG and IgM antibodies toIgE), asthma (often characterized, e.g., by IgG and IgM antibodies toIgE), inflammatory myopathies, and many other inflammatory,granulomatous, degenerative, and atrophic disorders.

In specific embodiments, antagonistic anti-neutrokine-alpha antibodiesare useful in the diagnosis and treatment or prevention of dilatedcardiomyopathy.

In a preferred embodiment, therapeutic and pharmaceutical compositionsof the invention, are used to treat, prevent, ameliorate, diagnose orprognose, a member of the group: autoimmune hemolytic anemia, as primaryglomerulonephritis, IgA glomerulonephritis, Goodpasture's syndrome,idiopathic thrombocytopenia, Multiple Sclerosis, Myasthenia Gravis,Pemphigus, polymyositis/dermatomyositis, relapsing polychondritis,rheumatoid arthritis, Sjögren's syndrome, systemic lupus erythematosus,Uveitis, vasculitis,and primary biliary cirrhosis.

In specific embodiments, pharmaceutical compositions of the inventionare useful in the diagnosis and treatment or prevention of liverdiseases, including but not limited to, primary biliary cirrhosis,primary sclerosing cholangitis, chronic hepatitis C infection,autoimmune hepatitis and alcoholic liver disease. In specificembodiments, antagonists of Neutrokine-alpha, such as ananti-Neutrokine-alpha specific antibody, are useful in the diagnosis andtreatment or prevention of liver diseases, including but not limited to,primary biliary cirrhosis, primary sclerosing cholangitis, chronichepatitis C infection, and alcoholic liver disease.

In another preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, an immune based-rheumatologic disease, such as,for example, SLE, rheumatoid arthritis, CREST syndrome (a variant ofschleroderma characterized by calcinosis, Raynaud's phenomenon,esophageal motility disorders, sclerodactyly, and telangiectasia.),Seronegative spondyloarthropathy (SpA), polymyositis/dermatomyositis,microscopic polyangiitis, hepatitis C-associated arthritis, Takayasu'sarteritis, and undifferentiated connective tissue disorder.

In a specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, rheumatoid arthritis and/or medical conditionsassociated therewith.

For example, an antibody, or antibodies, of the present invention areused to treat patients with clinical diagnosis of rheumatoid arthritis(RA). The patient treated preferably will not have a B cell malignancy.Moreover, the patient is optionally further treated with any one or moreagents employed for treating RA such as salicylate; nonsteroidalanti-inflammatory drugs such as indomethacin, phenylbutazone,phenylacetic acid derivatives (e.g. ibuprofen and fenoprofen),naphthalene acetic acids (naproxen), pyrrolealkanoic acid (tometin),indoleacetic acids (sulindac), halogenated anthranilic acid(meclofenamate sodium), piroxicam, zomepirac and diflunisal;antimalarials such as chloroquine; gold salts; penicillamine; orimmunosuppressive agents such as methotrexate or corticosteroids indosages known for such drugs or reduced dosages. Preferably however, thepatient is only treated with an antibody, or antibodies, of the presentinvention. Antibodies of the present invention are administered to theRA patient according to a dosing schedule as described infra, which maybe readily determined by one of ordinary skill in the art. The primaryresponse is determined by the Paulus index (Paulus et al. AthritisRheum. 33:477-484 (1990)), i.e. improvement in morning stiffness, numberof painful and inflamed joints, erythrocyte sedimentation (ESR), and atleast a 2-point improvement on a 5-point scale of disease severityassessed by patient and by physician. Administration of an antibody, orantibodies, of the present invention will alleviate one or more of thesymptoms of RA in the patient treated as described above.

In a specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, amelioate,diagnose or prognose, lupus and/or medical conditions associatedtherewith. Lupus-associated conditions that may be treated, prevented,ameliorated, prognosed and/or diagnosed with the antibodies and antibodycompositions of the invention include, but are not limited to,hematologic disorders (e.g., hemolytic anemia, leukopenia, lymphopenia,and thrombocytopenia), immunologic disorders (e.g., anti-DNA antibodies,and anti-Sm antibodies), rashes, photosensitivity, oral ulcers,arthritis, fever, fatigue, weight loss, serositis (e.g., pleuritus(pleurisy)), renal disorders (e.g., nephritis), neurological disorders(e.g., seizures, peripheral neuropathy, CNS related disorders),gastrointestinal disorders, Raynaud phenomenon, and pericarditis. In apreferred embodiment, therapeutic and pharmaceutical compositions of theinvention are used to treat, prevent, ameliorate, diagnose, or prognose,renal disorders associated with systemic lupus erythematosus. In a mostpreferred embodiment, therapeutic and pharmaceutical compositions of theinvention are used to treat, prevent, ameliorate, diagnose, or prognose,nephritis associated with systemic lupus erythematosus. In another mostpreferred embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent or amelioratelupus or glomerular nephritis.

In a further specific embodiment, antibodies of the invention are usedto treat, inhibit, prognose, diagnose or prevent hemolytic anemia. Forexample, patients diagnosed with autoimmune hemolytic anemia (AIHA),e.g., cryoglobinemia or Coombs positive anemia, are treated with anantibody, or antibodies, of the present invention. AIHA is an acquiredhemolytic anemia due to auto-antibodies that react with the patient'sred blood cells. The patient treated preferably will not have a B cellmalignancy. Further adjunct therapies (such as glucocorticoids,prednisone, azathioprine, cyclophosphamide, vinca-laden platelets orDanazol) may be combined with the antibody therapy, but preferably thepatient is treated with an antibody, or antibodies, of the presentinvention as a single-agent throughout the course of therapy. Antibodiesof the present invention are administered to the hemolytic anemiapatient according to a dosing schedule as described infra, which may bereadily determined by one of ordinary skill in the art. Overall responserate is determined based upon an improvement in blood counts, decreasedrequirement for transfusions, improved hemoglobin levels and/or adecrease in the evidence of hemolysis as determined by standard chemicalparameters. Administration of an antibody, or antibodies of the presentinvention will improve any one or more of the symptoms of hemolyticanemia in the patient treated as described above. For example, thepatient treated as described above will show an increase in hemoglobinand an improvement in chemical parameters of hemolysis or return tonormal as measured by serum lactic dehydrogenase and/or bilirubin.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, Sjögren's Syndrome and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, HIV infection and/or medical conditions associatedtherewith (e.g. AIDS).

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, Myasthenia gravis and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, IgA nephropathy and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, hemolytic anemia and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, thyroiditis and/or medical conditions associatedtherewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, Goodpasture's Syndrome and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, multiple sclerosis and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, chronic lymphocytic leukemia (CLL) and/or medicalconditions associated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, multiple myeloma and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, Non-Hodgkin's lymphoma and/or medical conditionsassociated therewith.

In another specific preferred embodiment, therapeutic and pharmaceuticalcompositions of the invention, are used to treat, prevent, ameliorate,diagnose or prognose, Hodgkin's disease and/or medical conditionsassociated therewith.

In another specific embodiment, antibodies of the invention are used totreat, inhibit, prognose, diagnose or prevent adult immunethrombocytopenic purpura. Adult immune thrombocytopenic purpura (ITP) isa relatively rare hematologic disorder that constitutes the most commonof the immune-mediated cytopenias. The disease typically presents withsevere thrombocytopenia that may be associated with acute hemorrhage inthe presence of normal to increased megakaryocytes in the bone marrow.Most patients with ITP have an IgG antibody directed against targetantigens on the outer surface of the platelet membrane, resulting inplatelet sequestration in the spleen and accelerated reticuloendothelialdestruction of platelets (Bussell, J. B. Hematol. Oncol. Clin. North Am.(4):179 (1990)). A number of therapeutic interventions have been shownto be effective in the treatment of ITP. Steroids are generallyconsidered first-line therapy, after which most patients are candidatesfor intravenous immunoglobulin (IVIG), splenectomy, or other medicaltherapies including vincristine or immunosuppressive/cytotoxic agents.Up to 80% of patients with ITP initially respond to a course ofsteroids, but far fewer have complete and lasting remissions.Splenectomy has been recommended as standard second-line therapy forsteroid failures, and leads to prolonged remission in nearly 60% ofcases yet may result in reduced immunity to infection. Splenectomy is amajor surgical procedure that may be associated with substantialmorbidity (15%) and mortality (2%). IVIG has also been used as secondline medical therapy, although only a small proportion of adult patientswith ITP achieve remission. Therapeutic options that would interferewith the production of autoantibodies by activated B cells without theassociated morbidities that occur with corticosteroids and/orsplenectomy would provide an important treatment approach for aproportion of patients with ITP. Patients with clinical diagnosis of ITPare treated with an antibody, or antibodies of the present invention,optionally in combination with steroid therapy. The patient treated willnot have a B cell malignancy. Antibodies of the present invention areadministered to the RA patient according to a dosing schedule asdescribed infra, which may be readily determined by one of ordinaryskill in the art. Overall patient response rate is determined based upona platelet count determined on two consecutive occasions two weeks apartfollowing treatments as described above. See, George et al. “IdiopathicThrombocytopenic Purpura: A Practice Guideline Developed by ExplicitMethods for The American Society of Hematology”, Blood 88:3-40 (1996),expressly incorporated herein by reference.

In another embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent or amelioratean IgE-mediated allergic reaction or histamine-mediated allergicreaction. Examples of allergic reactions include, but are not limitedto, asthma, rhinitis, eczema, chronic urticaria, and atopic dermatitis.In another embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent, or ameliorateanaphylaxis, hypersensitivity to an antigenic molecule, or blood groupincompatibility. In another embodiment, therapeutic or pharmaceuticalcompositions of the invention are administered to an animal to treat,prevent or ameliorate or modulate inflammation or an inflammatorydisorder. Examples of chronic and acute inflammatory disorders that maybe treated prevented or ameliorated with the therapeutic andpharmaceutical compositions of the invention include, but are notlimited to, chronic prostatitis, granulomatous prostatitis andmalacoplakia, inflammation associated with infection (e.g., septicshock, sepsis, or systemic inflammatory response syndrome (SIRS)),ischemia-reperfusion injury, endotoxin lethality, arthritis,complement-mediated hyperacute rejection, nephritis, cytokine orchemokine induced lung injury, Crohn's disease, inflammatory boweldisease, chronic and acute inflammatory pulmonary diseases, bacterialinfection, psoriasis, septicemia, cerebral malaria, arthritis,gastroenteritis, and glomerular nephritis.

In another embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent or ameliorateischemia and arteriosclerosis. Examples of such disorders include, butare not limited to, reperfusion damage (e.g., in the heart and/or brain)and cardiac hypertrophy.

Therapeutic or pharmaceutical compositions of the invention, may also beadministered to modulate blood clotting and to treat or prevent bloodclotting disorders, such as, for example, antibody-mediated thrombosis(i.e., antiphospholipid antibody syndrome (APS)). For example,therapeutic or pharmaceutical compositions of the invention, may inhibitthe proliferation and differentiation of cells involved in producinganticardiolipin antibodies. These compositions of the invention can beused to treat, prevent, ameliorate, diagnose, and/or prognose thromboticrelated events including, but not limited to, stroke (and recurrentstroke), heart attack, deep vein thrombosis, pulmonary embolism,myocardial infarction, coronary artery disease (e.g., antibody-mediatedcoronary artery disease), thrombosis, graft reocclusion followingcardiovascular surgery (e.g., coronary arterial bypass grafts, recurrentfetal loss, and recurrent cardiovascular thromboembolic events.

Therapeutic or pharmaceutical compositions of the invention, may also beadministered to treat, prevent, or ameliorate organ rejection orgraft-versus-host disease (GVHD) and/or conditions associated therewith.Organ rejection occurs by host immune cell destruction of thetransplanted tissue through an immune response. Similarly, an immuneresponse is also involved in GVHD, but, in this case, the foreigntransplanted immune cells destroy the host tissues. The administrationof antibodies of the invention, that inhibit an immune response, may bean effective therapy in preventing organ rejection or GVHD.

In another embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent or amelioratea disease or disorder diseases associated with increased apoptosisincluding, but not limited to, AIDS, neurodegenerative disorders (suchas Alzheimer's disease, Parkinson's disease, Amyotrophic lateralsclerosis, Retinitis pigmentosa, Cerebellar degeneration),myelodysplastic syndromes (such as aplastic anemia), ischemic injury(such as that caused by myocardial infarction, stroke and reperfusioninjury), toxin-induced liver disease (such as that caused by alcohol),septic shock, cachexia and anorexia. In another embodiment, therapeuticor pharmaceutical compositions of the invention are administered to ananimal to treat, prevent or ameliorate bone marrow failure, for example,aplastic anemia and myelodysplastic syndrome.

In another embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent or ameliorategrowth, progression, and/or metastases of malignancies and proliferativedisorders associated with increased cell survival, or the inhibition ofapoptosis. Examples of such disorders, include, but are not limited to,leukemia (e.g., acute leukemia such as acute lymphocytic leukemia andacute myelocytic leukemia), neoplasms, tumors (e.g., fibrosarcoma,myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma,angiosarcoma, endotheliosarcoma, lymphangiosarcoma,lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor,leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer,breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma,basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceousgland carcinoma, papillary carcinoma, papillary adenocarcinomas,cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renalcell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma,seminoma, embryonal carcinoma, Wilms' tumor, cervical cancer, testiculartumor, lung carcinoma, small cell lung carcinoma, bladder carcinoma,epithelial carcinoma, glioma, astrocytoma, medulloblastoma,craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acousticneuroma, oligodendroglioma, meningioma, melanoma, neuroblastoma, andretinoblastoma), heavy chain disease, metastases, or any disease ordisorder characterized by uncontrolled cell growth.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used to treat or prevent a disorder characterized byhpergammagloulinemia (e.g., AIDS, autoimmune diseases, and someimmunodeficiencies).

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used to treat or prevent a disorder characterized bydeficient serum immunoglobulin production, recurrent infections, and/orimmune system dysfunction. Moreover, therapeutic or pharmaceuticalcompositions of the invention may be used to treat or prevent infectionsof the joints, bones, skin, and/or parotid glands, blood-borneinfections (e.g., sepsis, meningitis, septic arthritis, and/orosteomyelitis), autoimmune diseases (e.g., those disclosed herein),inflammatory disorders, and malignancies, and/or any disease or disorderor condition associated with these infections, diseases, disordersand/or malignancies) including, but not limited to, CVID, other primaryimmune deficiencies, HIV disease, CLL, recurrent bronchitis, sinusitis,otitis media, conjunctivitis, pneumonia, hepatitis, meningitis, herpeszoster (e.g., severe herpes zoster), and/or pneumocystis carnii.

Therapeutic or pharmaceutical compositions of the invention of theinvention thereof, may be used to diagnose, prognose, treat or preventone or more of the following diseases or disorders, or conditionsassociated therewith: primary immunodeficiencies, immune-mediatedthrombocytopenia, Kawasaki syndrome, bone marrow transplant (e.g.,recent bone marrow transplant in adults or children), chronic B-celllymphocytic leukemia, HIV infection (e.g., adult or pediatric HIVinfection), chronic inflammatory demyelinating polyneuropathy, andpost-transfusion purpura.

Additionally, therapeutic or pharmaceutical compositions of theinvention may be used to diagnose, prognose, treat or prevent one ormore of the following diseases, disorders, or conditions associatedtherewith, Guillain-Barre syndrome, anemia (e.g., anemia associated withparvovirus B19, patients with stable multiple myeloma who are at highrisk for infection (e.g., recurrent infection), autoimmune hemolyticanemia (e.g., warm-type autoimmune hemolytic anemia), thrombocytopenia(e.g., neonatal thrombocytopenia), and immune-mediated neutropenia),transplantation (e.g., cytomegalovirus (CMV)-negative recipients ofCMV-positive organs), hypogammaglobulinemia (e.g., hypogammaglobulinemicneonates with risk factor for infection or morbidity), epilepsy (e.g.,intractable epilepsy), systemic vasculitic syndromes, myasthenia gravis(e.g., decompensation in myasthenia gravis), dermatomyositis, andpolymyositis.

Additional preferred embodiments of the invention include, but are notlimited to, the use of therapeutic or pharmaceutical compositions of theinvention in the following applications:

Administration to an animal (e.g., mouse, rat, rabbit, hamster, guineapig, pigs, micro-pig, chicken, camel, goat, horse, cow, sheep, dog, cat,non-human primate, and human, most preferably human) to boost the immunesystem to produce increased quantities of one or more antibodies (e.g.,IgG, IgA, IgM, and IgE), to induce higher affinity antibody production(e.g., IgG, IgA, IgM, and IgE), and/or to increase an immune response.In a specific nonexclusive embodiment, therapeutic or pharmaceuticalcompositions of the invention are administered to boost the immunesystem to produce increased quantities of IgG. In another specificnonexclusive embodiment, antibodies of the are administered to boost theimmune system to produce increased quantities of IgA. In anotherspecific nonexclusive embodiment antibodies of the invention areadministered to boost the immune system to produce increased quantitiesof IgM.

Administration to an animal (including, but not limited to, those listedabove, and also including transgenic animals) incapable of producingfunctional endogenous antibody molecules or having an otherwisecompromised endogenous immune system, but which is capable of producinghuman immunoglobulin molecules by means of a reconstituted or partiallyreconstituted immune system from another animal (see, e.g., publishedPCT Application Nos. WO98/24893, WO/9634096, WO/9633735, andWO/9110741).

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a vaccine adjuvant that enhances immuneresponsiveness to specific antigen. In a specific embodiment, thevaccine is an antibody described herein. In another specific embodiment,the vaccine adjuvant is a polynucleotide described herein (e.g., anantibody polynucleotide genetic vaccine adjuvant). As discussed herein,therapeutic or pharmaceutical compositions of the invention may beadministered using techniques known in the art, including but notlimited to, liposomal delivery, recombinant vector delivery, injectionof naked DNA, and gene gun delivery.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an adjuvant to enhance tumor-specific immuneresponses.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an adjuvant to enhance anti-viral immuneresponses. Anti-viral immune responses that may be enhanced using thecompositions of the invention as an adjuvant, include, but are notlimited to, virus and virus associated diseases or symptoms describedherein or otherwise known in the art. In specific embodiments, thecompositions of the invention are used as an adjuvant to enhance animmune response to a virus, disease, or symptom selected from the groupconsisting of: AIDS, meningitis, Dengue, EBV, and hepatitis (e.g.,hepatitis B). In another specific embodiment, the compositions of theinvention are used as an adjuvant to enhance an immune response to avirus, disease, or symptom selected from the group consisting of:HIV/AIDS, Respiratory syncytial virus, Dengue, Rotavirus, Japanese Bencephalitis, Influenza A and B, Parainfluenza, Measles,Cytomegalovirus, Rabies, Junin, Chikungunya, Rift Valley fever, Herpessimplex, and yellow fever. In another specific embodiment, thecompositions of the invention are used as an adjuvant to enhance animmune response to the HIV gp120 antigen.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an adjuvant to enhance anti-bacterial oranti-fungal immune responses. Anti-bacterial or anti-fungal immuneresponses that may be enhanced using the compositions of the inventionas an adjuvant, include bacteria or fungus and bacteria or fungusassociated diseases or symptoms described herein or otherwise known inthe art. In specific embodiments, the compositions of the invention areused as an adjuvant to enhance an immune response to a bacteria orfungus, disease, or symptom selected from the group consisting of:tetanus, Diphtheria, botulism, and meningitis type B. In anotherspecific embodiment, the compositions of the invention are used as anadjuvant to enhance an immune response to a bacteria or fungus, disease,or symptom selected from the group consisting of: Vibrio cholerae,Mycobacterium leprae, Salmonella typhi, Salmonella paratyphi, Neisseriameningitidis, Streptococcus pneumoniae, Group B streptococcus, Shigellaspp., Enterotoxigenic Escherichia coli, Enterohemorrhagic E. coli,Borrelia burgdorferi, and Plasmodium (malaria).

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an adjuvant to enhance anti-parasitic immuneresponses. Anti-parasitic immune responses that may be enhanced usingthe compositions of the invention as an adjuvant, include parasite andparasite associated diseases or symptoms described herein or otherwiseknown in the art. In specific embodiments, the compositions of theinvention are used as an adjuvant to enhance an immune response to aparasite. In another specific embodiment, the compositions of theinvention are used as an adjuvant to enhance an immune response toPlasmodium (malaria).

In a specific embodiment, compositions of the invention may beadministered to patients as vaccine adjuvants. In a further specificembodiment, compositions of the invention may be administered as vaccineadjuvants to patients suffering from an immune-deficiency. In a furtherspecific embodiment, compositions of the invention may be administeredas vaccine adjuvants to patients suffering from HIV.

In a specific embodiment, compositions of the invention may be used toincrease or enhance antigen-specific antibody responses to standard andexperimental vaccines. In a specific embodiment, compositions of theinvention may be used to enhance seroconversion in patients treated withstandard and experimental vaccines. In another specific embodiment,compositions of the invention may be used to increase the repertoire ofantibodies recognizing unique epitopes in response to standard andexperimental vaccination.

In a preferred embodiment, antibodies of the invention (includingantibody fragments and variants, and anti-antibody antibodies) increaseor enhance antigen-specific antibody responses to standard andexperimental vaccines by regulating binding of the soluble form of BLymphocyte Stimulator to a B Lymphocyte Stimulator receptor (e.g.,TACI—GenBank accession number AAC51790; BCMA—GenBank accession numberNP_(—)001183; and/or BAFF-R—GenBank accession number NP_(—)443177). Inanother preferred embodiment, antibodies of the invention (includingantibody fragments and variants, and anti-antibody antibodies) increaseor enhance antigen-specific antibody responses to standard andexperimental vaccines by regulating binding of the soluble form of APRILto an APRIL receptor (e.g., BCMA and TACI).

In a preferred embodiment, antibodies of the invention (includingantibody fragments and variants, and anti-antibody antibodies) increaseor enhance seroconversion in patients treated with standard andexperimental vaccines by regulating binding of the soluble form of BLymphocyte Stimulator to B Lymphocyte Stimulator receptor (e.g.,TACI—GenBank accession number AAC51790; BCMA—GenBank accession number NP001183; and/or BAFF-R—GenBank accession number NP_(—)443177). In anotherpreferred embodiment, antibodies of the invention (including antibodyfragments and variants, and anti-antibody antibodies) increase orenhance seroconversion in patients treated with standard andexperimental vaccines by regulating binding of the soluble form of APRILto an APRIL receptor (e.g., BCMA and TACI).

In a preferred embodiment, antibodies of the invention (includingantibody fragments and variants, and anti-antibody antibodies) increaseor enhance the repertoire of antibodies recognizing unique epitopes inresponse to standard and experimental vaccination by regulating bindingof the soluble form of B Lymphocyte Stimulator to a B LymphocyteStimulator receptor (e.g., TACI—GenBank accession number AAC51790;BCMA—GenBank accession number NP_(—)001183; and/or BAFF-R—GenBankaccession number NP_(—)443177). In another preferred embodiment,antibodies of the invention (including antibody fragments and variants,and anti-antibody antibodies) increase or enhance the repertoire ofantibodies recognizing unique epitopes in response to standard andexperimental vaccination by regulating binding of the soluble form ofAPRIL to an APRIL receptor (e.g., BCMA and TACI).

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a stimulator of B cell responsiveness topathogens.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent that elevates the immune status of anindividual prior to their receipt of immunosuppressive therapies.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent to induce higher affinity antibodies.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent to increase serum immunoglobulinconcentrations.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent to accelerate recovery ofimmunocompromised individuals.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent to boost immunoresponsiveness amongaged populations.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an immune system enhancer prior to, during, orafter bone marrow transplant and/or other transplants (e.g., allogeneicor xenogeneic organ transplantation). With respect to transplantation,compositions of the invention may be administered prior to, concomitantwith, and/or after transplantation. In a specific embodiment,compositions of the invention are administered after transplantation,prior to the beginning of recovery of T-cell populations. In anotherspecific embodiment, compositions of the invention are firstadministered after transplantation after the beginning of recovery of Tcell populations, but prior to full recovery of B cell populations.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent to boost immunoresponsiveness among Bcell immunodeficient individuals, such as, for example, an individualwho has undergone a partial or complete splenectomy. B cellimmunodeficiencies that may be ameliorated or treated by administeringthe antibodies and/or compositions of the invention include, but are notlimited to, severe combined immunodeficiency (SCID)-X linked,SCID-autosomal, adenosine deaminase deficiency (ADA deficiency),X-linked agammaglobulinemia (XLA), Bruton's disease, congenitalagammaglobulinemia, X-linked infantile agammaglobulinemia, acquiredagammaglobulinemia, adult onset agammaglobulinemia, late-onsetagammaglobulinemia, dysgammaglobulinemia, hypogammaglobulinemia,transient hypogammaglobulinemia of infancy, unspecifiedhypogammaglobulinemia, agammaglobulinemia, common variableimmunodeficiency (CVID) (acquired), Wiskott-Aldrich Syndrome (WAS),X-linked immunodeficiency with hyper IgM, non X-linked immunodeficiencywith hyper IgM, selective IgA deficiency, IgG subclass deficiency (withor without IgA deficiency), antibody deficiency with normal or elevatedIgs, immunodeficiency with thymoma, Ig heavy chain deletions, kappachain deficiency, B cell lymphoproliferative disorder (BLPD), selectiveIgM immunodeficiency, recessive agammaglobulinemia (Swiss type),reticular dysgenesis, neonatal neutropenia, severe congenitalleukopenia, thymic alymphoplasia-aplasia or dysplasia withimmunodeficiency, ataxia-telangiectasia, short limbed dwarfism, X-linkedlymphoproliferative syndrome (XLP), Nezelof syndrome-combinedimmunodeficiency with Igs, purine nucleoside phosphorylase deficiency(PNP), MHC Class II deficiency (Bare Lymphocyte Syndrome) and severecombined immunodeficiency.

In a specific embodiment, antibodies and/or compositions of theinvention are administered to treat or ameliorate selective IgAdeficiency.

In another specific embodiment, antibodies and/or compositions of theinvention are administered to treat or ameliorate ataxia-telangiectasia.

In another specific embodiment antibodies and/or compositions of theinvention are administered to treat or ameliorate common variableimmunodeficiency.

In another specific embodiment, antibodies and/or compositions of theinvention are administered to treat or ameliorate X-linkedagammaglobulinemia.

In another specific embodiment, antibodies and/or compositions of theinvention are administered to treat or ameliorate severe combinedimmunodeficiency (SCID).

In another specific embodiment, antibodies and/or compositions of theinvention are administered to treat or ameliorate Wiskott-Aldrichsyndrome.

In another specific embodiment, antibodies and/or compositions of theinvention are administered to treat or ameliorate X-linked Ig deficiencywith hyper IgM.

As an agent to boost immunoresponsiveness among individuals having anacquired loss of B cell function. Conditions resulting in an acquiredloss of B cell function that may be ameliorated or treated byadministering antibodies and/or compositions of the invention include,but are not limited to, HIV Infection, AIDS, bone marrow transplant, andB cell chronic lymphocytic leukemia (CLL).

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent to boost immunoresponsiveness amongindividuals having a temporary immune deficiency. Conditions resultingin a temporary immune deficiency that may be ameliorated or treated byadministering antibodies and/or compositions of the invention include,but are not limited to, recovery from viral infections (e.g.,influenza), conditions associated with malnutrition, recovery frominfectious mononucleosis, or conditions associated with stress, recoveryfrom measles, recovery from blood transfusion, recovery from surgery.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a regulator of antigen presentation bymonocytes, dendritic cells, T cells and/or B-cells. In one embodiment,antibody polypeptides or polynucleotides enhance antigen presentation orantagonize antigen presentation in vitro or in vivo. Moreover, inrelated embodiments, this enhancement or antagonization of antigenpresentation may be useful in anti-tumor treatment or to modulate theimmune system.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a mediator of mucosal immune responses. Theexpression of B Lymphocyte Stimulator on monocytes, the expression of BLymphocyte Stimulator receptor on B cells, and the responsiveness of Bcells to B Lymphocyte Stimulator suggests that it may be involved inexchange of signals between B cells and monocytes or theirdifferentiated progeny. This activity is in many ways analogous to theCD40-CD154 signaling between B cells and T cells. Anti-B LymphocyteStimulator antibodies and compositions of the invention may therefore begood regulators of T cell independent immune responses to environmentalpathogens. In particular, the unconventional B cell populations (CD5+)that are associated with mucosal sites and responsible for much of theinnate immunity in humans may respond to antibodies or compositions ofthe invention thereby enhancing or inhibiting individual's immunestatus.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an agent to direct an individual's immunesystem towards development of a humoral response (i.e. TH2) as opposedto a TH1 cellular response.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a means to induce tumor proliferation and thusmake it more susceptible to anti-neoplastic agents. For example,multiple myeloma is a slowly dividing disease and is thus refractory tovirtually all anti-neoplastic regimens. If these cells were forced toproliferate more rapidly, their susceptibility profile would likelychange.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a monocyte cell specific binding protein towhich specific activators or inhibitors of cell growth may be attached.The result would be to focus the activity of such activators orinhibitors onto normal, diseased, or neoplastic B cell populations.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a B cell specific binding protein to whichspecific activators or inhibitors of cell growth may be attached. Theresult would be to focus the activity of such activators or inhibitorsonto normal, diseased, or neoplastic B cell populations.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a means of detecting monocytic cells by virtueof its specificity. This application may require labeling the proteinwith biotin or other agents (e.g., as described herein) to afford ameans of detection.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a means of detecting B-lineage cells by virtueof its specificity. This application may require labeling the proteinwith biotin or other agents (e.g., as described herein) to afford ameans of detection.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a stimulator of B cell production inpathologies such as AIDS, chronic lymphocyte disorder and/or CommonVariable immunodeficiency.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as part of a monocyte selection device thefunction of which is to isolate monocytes from a heterogeneous mixtureof cell types. Antibodies of the invention could be coupled to a solidsupport to which monocytes would then specifically bind. Unbound cellswould be washed out and the bound cells subsequently eluted. Anon-limiting use of this selection would be to allow purging of tumorcells from, for example, bone marrow or peripheral blood prior totransplant.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as part of a B cell selection device the functionof which is to isolate B cells from a heterogeneous mixture of celltypes. Antibodies of the invention (that do not inhibit B LymphocyteStimulator/B Lymphocyte Stimulator Receptor interaction) binding solubleB Lymphocyte Stimulator could be coupled to a solid support to which Bcells would then specifically bind. Unbound cells would be washed outand the bound cells subsequently eluted. A non-limiting use of thisselection would be to allow purging of tumor cells from, for example,bone marrow or peripheral blood prior to transplant.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a therapy for generation and/or regenerationof lymphoid tissues following surgery, trauma or genetic defect.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a gene-based therapy for genetically inheriteddisorders resulting in immuno-incompetence such as observed among SCIDpatients.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as an antigen for the generation of antibodies toinhibit or enhance B Lymphocyte Stimulator mediated responses.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a means of activating monocytes/macrophages todefend against parasitic diseases that effect monocytes such asLeishmania.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as pretreatment of bone marrow samples prior totransplant. Such treatment would increase B cell representation and thusaccelerate recovery.

In a specific embodiment, therapeutic or pharmaceutical compositions ofthe invention are used as a means of regulating secreted cytokines thatare elicited by B Lymphocyte Stimulator and/or B Lymphocyte Stimulatorreceptor.

Antibody polypeptides or polynucleotides of the invention may be used tomodulate IgE concentrations in vitro or in vivo.

Additionally, antibody polypeptides or polynucleotides of the inventionmay be used to treat, prevent, and/or diagnose IgE-mediated allergicreactions. Such allergic reactions include, but are not limited to,asthma, rhinitis, and eczema.

In a specific embodiment, antibody polypeptides or polynucleotides ofthe invention, are administered to treat, prevent, diagnose, and/orameliorate selective IgA deficiency.

In another specific embodiment antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate ataxia-telangiectasia.

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate common variable immunodeficiency.

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate X-linked agammaglobulinemia.

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate severe combined immunodeficiency (SCID).

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate Wiskott-Aldrich syndrome.

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate X-linked Ig deficiency with hyper IgM. In a specificembodiment antibody polypeptides or polynucleotides of the invention areadministered to treat, prevent, diagnose, and/or ameliorate X-linked Igdeficiency with hyper IgM.

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, and/or diagnosechronic myelogenous leukemia, acute myelogenous leukemia, leukemia,hystiocytic leukemia, monocytic leukemia (e.g., acute monocyticleukemia), leukemic reticulosis, Shilling Type monocytic leukemia,and/or other leukemias derived from monocytes and/or monocytic cellsand/or tissues.

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate monocytic leukemoid reaction, as seen, for example, withtuberculosis.

In another specific embodiment, antibody polypeptides or polynucleotidesof the invention are administered to treat, prevent, diagnose, and/orameliorate monocytic leukocytosis, monocytic leukopenia, monocytopenia,and/or monocytosis.

In a specific embodiment, antibody polypeptides or polynucleotides ofthe invention are used to treat, prevent, detect, and/or diagnosemonocyte disorders and/or diseases, and/or conditions associatedtherewith.

In a specific embodiment, antibody polypeptides or polynucleotides ofthe invention are used to treat, prevent, detect, and/or diagnoseprimary B lymphocyte disorders and/or diseases, and/or conditionsassociated therewith. In one embodiment, such primary B lymphocytedisorders, diseases, and/or conditions are characterized by a completeor partial loss of humoral immunity. Primary B lymphocyte disorders,diseases, and/or conditions associated therewith that are characterizedby a complete or partial loss of humoral immunity and that may beprevented, treated, detected and/or diagnosed with compositions of theinvention include, but are not limited to, X-Linked Agammaglobulinemia(XLA), severe combined immunodeficiency disease (SCID), and selectiveIgA deficiency.

In a preferred embodiment antibody polypeptides or polynucleotides ofthe invention are used to treat, prevent, and/or diagnose diseases ordisorders affecting or conditions associated with any one or more of thevarious mucous membranes of the body. Such diseases or disordersinclude, but are not limited to, for example, mucositis, mucoclasis,mucocolitis, mucocutaneous leishmaniasis (such as, for example, Americanleishmaniasis, leishmaniasis americana, nasopharyngeal leishmaniasis,and New World leishmaniasis), mucocutaneous lymph node syndrome (forexample, Kawasaki disease), mucoenteritis, mucoepidermoid carcinoma,mucoepidermoid tumor, mucoepithelial dysplasia, mucoid adenocarcinoma,mucoid degeneration, myxoid degeneration; myxomatous degeneration;myxomatosis, mucoid medial degeneration (for example, cystic medialnecrosis), mucolipidosis (including, for example, mucolipidosis I,mucolipidosis II, mucolipidosis III, and mucolipidosis IV), mucolysisdisorders, mucomembranous enteritis, mucoenteritis,mucopolysaccharidosis (such as, for example, type Imucopolysaccharidosis (i.e., Hurler's syndrome), type ISmucopolysaccharidosis (i.e., Scheie's syndrome or type Vmucopolysaccharidosis), type II mucopolysaccharidosis (i.e., Hunter'ssyndrome), type III mucopolysaccharidosis (i.e., Sanfilippo's syndrome),type IV mucopolysaccharidosis (i.e., Morquio's syndrome), type VImucopolysaccharidosis (i.e., Maroteaux-Lamy syndrome), type VIImucopolysaccharidosis (i.e, mucopolysaccharidosis due tobeta-glucuronidase deficiency), and mucosulfatidosis),mucopolysacchariduria, mucopurulent conjunctivitis, mucopus,mucormycosis (i.e., zygomycosis), mucosal disease (i.e., bovine virusdiarrhea), mucous colitis (such as, for example, mucocolitis andmyxomembranous colitis), and mucoviscidosis (such as, for example,cystic fibrosis, cystic fibrosis of the pancreas, Clarke-Hadfieldsyndrome, fibrocystic disease of the pancreas, mucoviscidosis, andviscidosis). In a highly preferred embodiment, antibody polypeptides orpolynucleotides of the invention are used to treat, prevent, and/ordiagnose mucositis, especially as associated with chemotherapy.

In a preferred embodiment, antibody polypeptides or polynucleotides ofthe invention are used to treat, prevent, and/or diagnose diseases ordisorders affecting or conditions associated with sinusitis.

An additional condition, disease or symptom that can be treated,prevented, and/or diagnosed by antibody polypeptides or polynucleotidesof the invention is osteomyelitis.

An additional condition, disease or symptom that can be treated,prevented, and/or diagnosed by antibody polypeptides or polynucleotidesof the invention is endocarditis.

All of the above described applications as they may apply to veterinarymedicine.

Antibody polypeptides or polynucleotides of the invention may be used totreat, prevent, and/or diagnose diseases and disorders of the pulmonarysystem (e.g., bronchi such as, for example, sinopulmonary and bronchialinfections and conditions associated with such diseases and disordersand other respiratory diseases and disorders. In specific embodiments,such diseases and disorders include, but are not limited to, bronchialadenoma, bronchial asthma, pneumonia (such as, e.g., bronchialpneumonia, bronchopneumonia, and tuberculous bronchopneumonia), chronicobstructive pulmonary disease (COPD), bronchial polyps, bronchiectasia(such as, e.g., bronchiectasia sicca, cylindrical bronchiectasis, andsaccular bronchiectasis), bronchiolar adenocarcinoma, bronchiolarcarcinoma, bronchiolitis (such as, e.g., exudative bronchiolitis,bronchiolitis fibrosa obliterans, and proliferative bronchiolitis),bronchiolo-alveolar carcinoma, bronchitic asthma, bronchitis (such as,e.g., asthmatic bronchitis, Castellani's bronchitis, chronic bronchitis,croupous bronchitis, fibrinous bronchitis, hemorrhagic bronchitis,infectious avian bronchitis, obliterative bronchitis, plasticbronchitis, pseudomembranous bronchitis, putrid bronchitis, andverminous bronchitis), bronchocentric granulomatosis, bronchoedema,bronchoesophageal fistula, bronchogenic carcinoma, bronchogenic cyst,broncholithiasis, bronchomalacia, bronchomycosis (such as, e.g.,bronchopulmonary aspergillosis), bronchopulmonary spirochetosis,hemorrhagic bronchitis, bronchorrhea, bronchospasm, bronchostaxis,bronchostenosis, Biot's respiration, bronchial respiration, Kussmaulrespiration, Kussmaul-Kien respiration, respiratory acidosis,respiratory alkalosis, respiratory distress syndrome of the newborn,respiratory insufficiency, respiratory scleroma, respiratory syncytialvirus, and the like.

In a specific embodiment, antibody polypeptides or polynucleotides ofthe invention are used to treat, prevent, and/or diagnose chronicobstructive pulmonary disease (COPD).

In another embodiment, antibody polypeptides or polynucleotides of theinvention are used to treat, prevent, and/or diagnose fibroses andconditions associated with fibroses, including, but not limited to,cystic fibrosis (including such fibroses as cystic fibrosis of thepancreas, Clarke-Hadfield syndrome, fibrocystic disease of the pancreas,mucoviscidosis, and viscidosis), endomyocardial fibrosis, idiopathicretroperitoneal fibrosis, leptomeningeal fibrosis, mediastinal fibrosis,nodular subepidermal fibrosis, pericentral fibrosis, perimuscularfibrosis, pipestem fibrosis, replacement fibrosis, subadventitialfibrosis, and Symmers' clay pipestem fibrosis.

In another embodiment, therapeutic or pharmaceutical compositions of theinvention are administered to an animal to treat, prevent or ameliorateinfectious diseases. Infectious diseases include diseases associatedwith yeast, fungal, viral and bacterial infections. Viruses causingviral infections which can be treated or prevented in accordance withthis invention include, but are not limited to, retroviruses (e.g.,human T-cell lymphotrophic virus (HTLV) types I and II and humanimmunodeficiency virus (HIV)), herpes viruses (e.g., herpes simplexvirus (HSV) types I and II, Epstein-Barr virus, HHV6-HHV8, andcytomegalovirus), arenavirues (e.g., lassa fever virus), paramyxoviruses(e.g., morbillivirus virus, human respiratory syncytial virus, mumps,and pneumovirus), adenoviruses, bunyaviruses (e.g., hantavirus),cornaviruses, filoviruses (e.g., Ebola virus), flaviviruses (e.g.,hepatitis C virus (HCV), yellow fever virus, and Japanese encephalitisvirus), hepadnaviruses (e.g., hepatitis B viruses (HBV)),orthomyoviruses (e.g., influenza viruses A, B and C), papovaviruses(e.g., papillomavirues), picornaviruses (e.g., rhinoviruses,enteroviruses and hepatitis A viruses), poxviruses, reoviruses (e.g.,rotavirues), togaviruses (e.g., rubella virus), rhabdoviruses (e.g.,rabies virus). Microbial pathogens causing bacterial infections include,but are not limited to, Streptococcus pyogenes, Streptococcuspneumoniae, Neisseria gonorrhoea, Neisseria meningitidis,Corynebacterium diphtheriae, Clostridium botulinum, Clostridiumperfringens, Clostridium tetani, Haemophilus influenzae, Klebsiellapneumoniae, Klebsiella ozaenae, Klebsiella rhinoscleromotis,Staphylococcus aureus, Vibrio cholerae, Escherichia coli, Pseudomonasaeruginosa, Campylobacter (Vibrio) fetus, Campylobacter jejuni,Aeromonas hydrophila, Bacillus cereus, Edwardsiella tarda, Yersiniaenterocolitica, Yersinia pestis, Yersinia pseudotuberculosis, Shigelladysenteriae, Shigella flexneri, Shigella sonnei, Salmonella typhimurium,Treponema pallidum, Treponema pertenue, Treponema carateneum, Borreliavincentii, Borrelia burgdorferi, Leptospira icterohemorrhagiae,Mycobacterium tuberculosis, Toxoplasma gondii, Pneumocystis carinii,Francisella tularensis, Brucella abortus, Brucella suis, Brucellamelitensis, Mycoplasma spp., Rickettsia prowazeki, Rickettsiatsutsugumushi, Chlamydia spp., and Helicobacter pylori.

Gene Therapy

In a specific embodiment, nucleic acids comprising sequences encodingantibodies or functional derivatives thereof, are administered to treat,inhibit or prevent a disease or disorder associated with aberrantexpression and/or activity of B Lymphocyte Stimulator and/or itsreceptor, by way of gene therapy. Gene therapy refers to therapyperformed by the administration to a subject of an expressed orexpressible nucleic acid. In this embodiment of the invention, thenucleic acids produce their encoded protein that mediates a therapeuticeffect.

Any of the methods for gene therapy available in the art can be usedaccording to the present invention. Exemplary methods are describedbelow.

For general reviews of the methods of gene therapy, see Goldspiel etal., Clinical Pharmacy 12:488-505 (1993); Wu and Wu, Biotherapy 3:87-95(1991); Tolstoshev, Ann. Rev. Pharmacol. Toxicol. 32:573-596 (1993);Mulligan, Science 260:926-932 (1993); and Morgan and Anderson, Ann. Rev.Biochem. 62:191-217 (1993); May, TIBTECH 1 l(5):155-215 (1993). Methodscommonly known in the art of recombinant DNA technology which can beused are described in Ausubel et al. (eds.), Current Protocols inMolecular Biology, John Wiley & Sons, NY (1993); and Kriegler, GeneTransfer and Expression, A Laboratory Manual, Stockton Press, NY (1990).

In a preferred aspect, a composition of the invention comprises, oralternatively consists of, nucleic acids encoding an antibody, saidnucleic acids being part of an expression vector that expresses theantibody or fragments or chimeric proteins or heavy or light chainsthereof in a suitable host. In particular, such nucleic acids havepromoters, preferably heterologous promoters, operably linked to theantibody coding region, said promoter being inducible or constitutive,and, optionally, tissue-specific. In another particular embodiment,nucleic acid molecules are used in which the antibody coding sequencesand any other desired sequences are flanked by regions that promotehomologous recombination at a desired site in the genome, thus providingfor intrachromosomal expression of the antibody encoding nucleic acids(Koller and Smithies, Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989);Zijlstra et al., Nature 342:435-438 (1989). In specific embodiments, theexpressed antibody molecule is an scFv; alternatively, the nucleic acidsequences include sequences encoding both the heavy and light chains, orfragments or variants thereof, of an antibody.

Delivery of the nucleic acids into a patient may be either direct, inwhich case the patient is directly exposed to the nucleic acid ornucleic acid-carrying vectors, or indirect, in which case, cells arefirst transformed with the nucleic acids in vitro, then transplantedinto the patient. These two approaches are known, respectively, as invivo or ex vivo gene therapy.

In a specific embodiment, the nucleic acid sequences are directlyadministered in vivo, where it is expressed to produce the encodedproduct. This can be accomplished by any of numerous methods known inthe art, e.g., by constructing them as part of an appropriate nucleicacid expression vector and administering it so that they becomeintracellular, e.g., by infection using defective or attenuatedretrovirals or other viral vectors (see U.S. Pat. No. 4,980,286), or bydirect injection of naked DNA, or by use of microparticle bombardment(e.g., a gene gun; Biolistic, Dupont), or coating with lipids orcell-surface receptors or transfecting agents, encapsulation inliposomes, microparticles, or microcapsules, or by administering them inlinkage to a peptide which is known to enter the nucleus, byadministering it in linkage to a ligand subject to receptor-mediatedendocytosis (see, e.g., Wu and Wu, J. Biol. Chem. 262:4429-4432 (1987))(which can be used to target cell types specifically expressing thereceptors), etc. In another embodiment, nucleic acid-ligand complexescan be formed in which the ligand comprises a fusogenic viral peptide todisrupt endosomes, allowing the nucleic acid to avoid lysosomaldegradation. In yet another embodiment, the nucleic acid can be targetedin vivo for cell specific uptake and expression, by targeting a specificreceptor (see, e.g., PCT Publications WO 92/06 180; WO 92/22635;WO92/203 16; WO93/14188, WO 93/20221). Alternatively, the nucleic acidcan be introduced intracellularly and incorporated within host cell DNAfor expression, by homologous recombination (Koller and Smithies, Proc.Natl. Acad. Sci. USA 86:8932-8935 (1989); Zijlstra et al., Nature342:435-438 (1989)).

In a specific embodiment, viral vectors that contains nucleic acidsequences encoding an antibody of the invention or fragments or variantsthereof are used. For example, a retroviral vector can be used (seeMiller et al., Meth. Enzymol. 217:581-599 (1993)). These retroviralvectors contain the components necessary for the correct packaging ofthe viral genome and integration into the host cell DNA. The nucleicacid sequences encoding the antibody to be used in gene therapy arecloned into one or more vectors, which facilitates delivery of the geneinto a patient. More detail about retroviral vectors can be found inBoesen et al., Biotherapy 6:29 1-302 (1994), which describes the use ofa retroviral vector to deliver the mdr 1 gene to hematopoietic stemcells in order to make the stem cells more resistant to chemotherapy.Other references illustrating the use of retroviral vectors in genetherapy are: Clowes et al., J. Clin. Invest. 93:644-651(1994); Klein etal., Blood 83:1467-1473 (1994); Salmons and Gunzberg, Human Gene Therapy4:129-141 (1993); and Grossman and Wilson, Curr. Opin. in Genetics andDevel. 3:110-114 (1993).

Adenoviruses are other viral vectors that can be used in gene therapy.Adenoviruses are especially attractive vehicles for delivering genes torespiratory epithelia. Adenoviruses naturally infect respiratoryepithelia where they cause a mild disease. Other targets foradenovirus-based delivery systems are liver, the central nervous system,endothelial cells, and muscle. Adenoviruses have the advantage of beingcapable of infecting non-dividing cells. Kozarsky and Wilson, CurrentOpinion in Genetics and Development 3:499-503 (1993) present a review ofadenovirus-based gene therapy. Bout et al., Human Gene Therapy 5:3-10(1994) demonstrated the use of adenovirus vectors to transfer genes tothe respiratory epithelia of rhesus monkeys. Other instances of the useof adenoviruses in gene therapy can be found in Rosenfeld et al.,Science 252:431-434 (1991); Rosenfeld et al., Cell 68:143-155 (1992);Mastrangeli et al., J. Clin. Invest. 91:225-234 (1993); PCT PublicationWO94/12649; and Wang, et al., Gene Therapy 2:775-783 (1995). In apreferred embodiment, adenovirus vectors are used.

Adeno-associated virus (AAV) has also been proposed for use in genetherapy (Walsh et al., Proc. Soc. Exp. Biol. Med. 204:289-300 (1993);U.S. Pat. No. 5,436,146).

Another approach to gene therapy involves transferring a gene to cellsin tissue culture by such methods as electroporation, lipofection,calcium phosphate mediated transfection, or viral infection. Usually,the method of transfer includes the transfer of a selectable marker tothe cells. The cells are then placed under selection to isolate thosecells that have taken up and are expressing the transferred gene. Thosecells are then delivered to a patient.

In this embodiment, the nucleic acid is introduced into a cell prior toadministration in vivo of the resulting recombinant cell. Suchintroduction can be carried out by any method known in the art,including but not limited to transfection, electroporation,microinjection, infection with a viral or bacteriophage vectorcontaining the nucleic acid sequences, cell fusion, chromosome-mediatedgene transfer, microcell mediated gene transfer, spheroplast fusion,etc. Numerous techniques are known in the art for the introduction offoreign genes into cells (see, e.g., Loeffler and Behr, Meth. Enzymol.217:599-618 (1993); Cohen et al., Meth. Enzymol. 217:618-644 (1993);Clin. Pharma. Ther. 29:69-92m (1985)) and may be used in accordance withthe present invention, provided that the necessary developmental andphysiological functions of the recipient cells are not disrupted. Thetechnique should provide for the stable transfer of the nucleic acid tothe cell, so that the nucleic acid is expressible by the cell andpreferably heritable and expressible by its cell progeny.

The resulting recombinant cells can be delivered to a patient by variousmethods known in the art. Recombinant blood cells (e.g., hematopoieticstem or progenitor cells) are preferably administered intravenously. Theamount of cells envisioned for use depends on the desired effect,patient state, etc., and can be determined by one skilled in the art.

Cells into which a nucleic acid can be introduced for purposes of genetherapy encompass any desired, available cell type, and include but arenot limited to epithelial cells, endothelial cells, keratinocytes,fibroblasts, muscle cells, hepatocytes; blood cells such as Tlymphocytes, B lymphocytes, monocytes, macrophages, neutrophils,eosinophils, megakaryocytes, granulocytes; various stem or progenitorcells, in particular hematopoietic stem or progenitor cells, e.g., asobtained from bone marrow, umbilical cord blood, peripheral blood, fetalliver, etc.

In a preferred embodiment, the cell used for gene therapy is autologousto the patient.

In an embodiment in which recombinant cells are used in gene therapy,nucleic acid sequences encoding an antibody or fragment thereof areintroduced into the cells such that they are expressible by the cells ortheir progeny, and the recombinant cells are then administered in vivofor therapeutic effect. In a specific embodiment, stem or progenitorcells are used. Any stem and/or progenitor cells which can be isolatedand maintained in vitro can potentially be used in accordance with thisembodiment of the present invention (see e.g. PCT Publication WO94/08598; Stemple and Anderson, Cell 7 1:973-985 (1992); Rheinwald,Meth. Cell Bio. 21A:229 (1980); and Pittelkow and Scott, Mayo ClinicProc. 61:771 (1986)).

In a specific embodiment, the nucleic acid to be introduced for purposesof gene therapy comprises an inducible promoter operably linked to thecoding region, such that expression of the nucleic acid is controllableby controlling the presence or absence of the appropriate inducer oftranscription.

Demonstration of Therapeutic or Prophylactic Utility of a Composition

The compounds of the invention are preferably tested in vitro, and thenin vivo for the desired therapeutic or prophylactic activity, prior touse in humans. For example, in vitro assays which can be used todetermine whether administration of a specific antibody or compositionof the present invention is indicated, include in vitro cell cultureassays in which a patient tissue sample is grown in culture, and exposedto or otherwise administered an antibody or composition of the presentinvention, and the effect of such an antibody or composition of thepresent invention upon the tissue sample is observed. In variousspecific embodiments, in vitro assays can be carried out withrepresentative cells of cell types involved in a patient's disorder, todetermine if an antibody or composition of the present invention has adesired effect upon such cell types. Preferably, the antibodies orcompositions of the invention are also tested in in vitro assays andanimal model systems prior to administration to humans.

Antibodies or compositions of the present invention for use in therapycan be tested for their toxicity in suitable animal model systems,including but not limited to rats, mice, chicken, cows, monkeys, andrabbits. For in vivo testing of an antibody or composition's toxicityany animal model system known in the art may be used.

Efficacy in treating or preventing viral infection may be demonstratedby detecting the ability of an antibody or composition of the inventionto inhibit the replication of the virus, to inhibit transmission orprevent the virus from establishing itself in its host, or to prevent,ameliorate or alleviate the symptoms of disease a progression. Thetreatment is considered therapeutic if there is, for example, areduction in viral load, amelioration of one or more symptoms, or adecrease in mortality and/or morbidity following administration of anantibody or composition of the invention.

Antibodies or compositions of the invention can be tested for theability to induce the expression of cytokines such as IFN-γ, bycontacting cells, preferably human cells, with an antibody orcomposition of the invention or a control antibody or controlcomposition and determining the ability of the antibody or compositionof the invention to induce one or more cytokines. Techniques known tothose of skill in the art can be used to measure the level of expressionof cytokines. For example, the level of expression of cytokines can bemeasured by analyzing the level of RNA of cytokines by, for example,RT-PCR and Northern blot analysis, and by analyzing the level ofcytokines by, for example, immunoprecipitation followed by western blotanalysis and ELISA. In a preferred embodiment, a compound of theinvention is tested for its ability to induce the expression of IFN-γ.

Antibodies or compositions of the invention can be tested for theirability to modulate the biological activity of immune cells bycontacting immune cells, preferably human immune cells (e.g., T-cells,B-cells, and Natural Killer cells), with an antibody or composition ofthe invention or a control compound and determining the ability of theantibody or composition of the invention to modulate (i.e, increase ordecrease) the biological activity of immune cells. The ability of anantibody or composition of the invention to modulate the biologicalactivity of immune cells can be assessed by detecting the expression ofantigens, detecting the proliferation of immune cells (i.e., B-cellproliferation), detecting the activation of signaling molecules,detecting the effector function of immune cells, or detecting thedifferentiation of immune cells. Techniques known to those of skill inthe art can be used for measuring these activities. For example,cellular proliferation can be assayed by ³H-thymidine incorporationassays and trypan blue cell counts. Antigen expression can be assayed,for example, by immunoassays including, but not limited to, competitiveand non-competitive assay systems using techniques such as westernblots, immunohistochemistry radioimmunoassays, ELISA (enzyme linkedimmunosorbent assay), “sandwich” immunoassays, immunoprecipitationassays, precipitant reactions, gel diffusion precipitant reactions,immunodiffusion assays, agglutination assays, complement-fixationassays, immunoradiometric assays, fluorescent immunoassays, protein Aimmunoassays and FACS analysis. The activation of signaling moleculescan be assayed, for example, by kinase assays and electrophoretic shiftassays (EMSAs). In a preferred embodiment, the ability of an antibody orcomposition of the invention to induce B-cell proliferation is measured.In another preferred embodiment, the ability of an antibody orcomposition of the invention to modulate immunoglobulin expression ismeasured.

Antibodies or compositions of the invention can be tested for theirability to reduce tumor formation in in vitro, ex vivo and in vivoassays. Antibodies or compositions of the invention can also be testedfor their ability to inhibit viral replication or reduce viral load inin vitro and in vivo assays. Antibodies or compositions of the inventioncan also be tested for their ability to reduce bacterial numbers in invitro and in vivo assays known to those of skill in the art. Antibodiesor compositions of the invention can also be tested for their ability toalleviate of one or more symptoms associated with cancer, an immunedisorder (e.g., an inflammatory disease), a neurological disorder or aninfectious disease. Antibodies or compositions of the invention can alsobe tested for their ability to decrease the time course of theinfectious disease. Further, antibodies or compositions of the inventioncan be tested for their ability to increase the survival period ofanimals suffering from disease or disorder, including cancer, an immunedisorder or an infectious disease. Techniques known to those of skill inthe art can be used to analyze the function of the antibodies orcompositions of the invention in vivo.

Therapeutic/Prophylactic Compositions and Administration

The invention provides methods of treatment, inhibition and prophylaxisby administration to a subject of an effective amount of antibody (orfragment or variant thereof) or pharmaceutical composition of theinvention, preferably an antibody of the invention. In a preferredaspect, an antibody or fragment or variant thereof is substantiallypurified (i.e., substantially free from substances that limit its effector produce undesired side-effects). The subject is preferably an animal,including but not limited to, animals such as cows, pigs, horses,chickens, cats, dogs, etc., and is preferably a mammal, and mostpreferably a human.

Formulations and methods of administration that can be employed when thecompound comprises a nucleic acid or an immunoglobulin as describedabove; additional appropriate formulations and routes of administrationcan be selected from among those described herein below.

Various delivery systems are known and can be used to administerantibody or fragment or variant thereof of the invention, e.g.,encapsulation in liposomes, microparticles, microcapsules, recombinantcells capable of expressing the antibody or antibody fragment,receptor-mediated endocytosis (see, e.g., Wu and Wu, J. Biol. Chem.262:4429-4432 (1987)), construction of a nucleic acid as part of aretroviral or other vector, etc. Methods of introduction include, butare not limited to, intradermal, intramuscular, intraperitoneal,intravenous, subcutaneous, intranasal, epidural, and oral routes. Thecompositions may be administered by any convenient route, for example byinfusion or bolus injection, by absorption through epithelial ormucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa,etc.) and may be administered together with other biologically activeagents. Administration can be systemic or local. In addition, it may bedesirable to introduce the pharmaceutical compositions of the inventioninto the central nervous system by any suitable route, includingintraventricular and intrathecal injection; intraventricular injectionmay be facilitated by an intraventricular catheter, for example,attached to a reservoir, such as an Ommaya reservoir. Pulmonaryadministration can also be employed, e.g., by use of an inhaler ornebulizer, and formulation with an aerosolizing agent.

In a preferred embodiment the antibody of the invention is formulated in10 mM sodium citrate, 1.9% glycine, 0.5% sucrose, 0.01% polysorbate 80,pH 6.5 (±0.3). In another preferred embodiment, the antibody of theinvention is formulated in 10 mM sodium citrate, 1.9% glycine, 0.5%sucrose, 0.01% polysorbate 80, pH 6.5 (±0.3) for intravenousadministration.

In a preferred embodiment the antibody of the invention is formulated in110 mM sodium citrate, 8% sucrose, 0.04% (w/v) polysorbate 80 (pH 6.5)(±0.3). In another preferred embodiment, the antibody of the inventionis formulated in 10 mM sodium citrate, 8% sucrose, 0.04% (w/v)polysorbate 80 (pH 6.5) for intravenous administration. In anotherpreferred embodiment, the antibody of the invention is formulated in 10mM sodium citrate, 8% sucrose, 0.04% (w/v) polysorbate 80 (pH 6.5) forsubcutaneous administration.

In a specific embodiment, it may be desirable to administer thepharmaceutical compositions of the invention locally to the area in needof treatment; this may be achieved by, for example, and not by way oflimitation, local infusion during surgery, topical application, e.g., inconjunction with a wound dressing after surgery, by injection, by meansof a catheter, by means of a suppository, or by means of an implant,said implant being of a porous, non-porous, or gelatinous material,including membranes, such as sialastic membranes, or fibers. Preferably,when administering a protein, including an antibody, of the invention,care must be taken to use materials to which the protein does notabsorb.

In another embodiment, the composition can be delivered in a vesicle, inparticular a liposome (see Langer, Science 249:1527-1533 (1990); Treatet al., in Liposomes in the Therapy of Infectious Disease and Cancer,Lopez-Berestein and Fidler (eds.), Liss, New York, pp. 353-365 (1989);Lopez-Berestein, ibid., pp. 317-327; see generally ibid.).

In yet another embodiment, the composition can be delivered in acontrolled release system. In one embodiment, a pump may be used (seeLanger, supra; Sefton, CRC Crit. Ref. Biomed. Eng. 14:20 1 (1987);Buchwald et al., Surgery 88:507 (1980); Saudek et al., N. Engl. J. Med.321:574 (1989)). In another embodiment, polymeric materials can be used(see Medical Applications of Controlled Release, Langer and Wise (eds.),CRC Pres., Boca Raton, Fla. (1974); Controlled Drug Bioavailability,Drug Product Design and Performance, Smolen and Ball (eds.), Wiley, NewYork (1984); Ranger and Peppas, J., Macromol. Sci. Rev. Macromol. Chem.23:61 (1983); see also Levy et al., Science 228:190 (1985); During etal., Ann. Neurol. 25:35 1 (1989); Howard et al., J. Neurosurg. 7 1:105(1989)). In yet another embodiment, a controlled release system can beplaced in proximity of the therapeutic target, i.e., the brain, thusrequiring only a fraction of the systemic dose (see, e.g., Goodson, inMedical Applications of Controlled Release, supra, vol. 2, pp. 115-138(1984)).

Other controlled release systems are discussed in the review by Langer(Science 249:1527-1533 (1990)).

In a specific embodiment where the composition of the invention is anucleic acid encoding a protein, the nucleic acid can be administered invivo to promote expression of its encoded protein, by constructing it aspart of an appropriate nucleic acid expression vector and administeringit so that it becomes intracellular, e.g., by use of a retroviral vector(see U.S. Pat. No. 4,980,286), or by direct injection, or by use ofmicroparticle bombardment (e.g., a gene gun; Biolistic, Dupont), orcoating with lipids or cell-surface receptors or transfecting agents, orby administering it in linkage to a homeobox-like peptide which is knownto enter the nucleus (see e.g., Joliot et al., Proc. Natl. Acad. Sci.USA 88:1864-1868 (1991)), etc. Alternatively, a nucleic acid can beintroduced intracellularly and incorporated within host cell DNA forexpression, by homologous recombination.

The present invention also provides pharmaceutical compositions. Suchcompositions comprise a therapeutically effective amount of an antibodyor a fragment thereof, and a pharmaceutically acceptable carrier. In aspecific embodiment, the term “pharmaceutically acceptable” meansapproved by a regulatory agency of the Federal or a state government orlisted in the U.S. Pharmacopeia or other generally recognizedpharmacopeia for use in animals, and more particularly in humans. Theterm “carrier” refers to a diluent, adjuvant, excipient, or vehicle withwhich the therapeutic is administered. Such pharmaceutical carriers canbe sterile liquids, such as water and oils, including those ofpetroleum, animal, vegetable or synthetic origin, such as peanut oil,soybean oil, mineral oil, sesame oil and the like. Water is a preferredcarrier when the pharmaceutical composition is administeredintravenously. Saline solutions and aqueous dextrose and glycerolsolutions can also be employed as liquid carriers, particularly forinjectable solutions. Suitable pharmaceutical excipients include starch,glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silicagel, sodium stearate, glycerol monostearate, talc, sodium chloride,dried skim milk, glycerol, propylene, glycol, water, ethanol and thelike. The composition, if desired, can also contain minor amounts ofwetting or emulsifying agents, or pH buffering agents. Thesecompositions can take the form of solutions, suspensions, emulsion,tablets, pills, capsules, powders, sustained-release formulations andthe like. The composition can be formulated as a suppository, withtraditional binders and carriers such as triglycerides. Oral formulationcan include standard carriers such as pharmaceutical grades of mannitol,lactose, starch, magnesium stearate, sodium saccharine, cellulose,magnesium carbonate, etc. Examples of suitable pharmaceutical carriersare described in “Remington's Pharmaceutical Sciences” by E. W. Martin.Such compositions will contain a therapeutically effective amount of theantibody or fragment thereof, preferably in purified form, together witha suitable amount of carrier so as to provide the form for properadministration to the patient. The formulation should suit the mode ofadministration.

In a preferred embodiment, the composition is formulated in accordancewith routine procedures as a pharmaceutical composition adapted forintravenous administration to human beings. Typically, compositions forintravenous administration are solutions in sterile isotonic aqueousbuffer. Where necessary, the composition may also include a solubilizingagent and a local anesthetic such as lignocamne to ease pain at the siteof the injection. Generally, the ingredients are supplied eitherseparately or mixed together in unit dosage form, for example, as a drylyophilized powder or water free concentrate in a hermetically sealedcontainer such as an ampoule or sachette indicating the quantity ofactive agent. Where the composition is to be administered by infusion,it can be dispensed with an infusion bottle containing sterilepharmaceutical grade water or saline. Where the composition isadministered by injection, an ampoule of sterile water for injection orsaline can be provided so that the ingredients may be mixed prior toadministration.

The compositions of the invention can be formulated as neutral or saltforms. Pharmaceutically acceptable salts include those formed withanions such as those derived from hydrochloric, phosphoric, acetic,oxalic, tartaric acids, etc., and those formed with cations such asthose derived from sodium, potassium, ammonium, calcium, ferrichydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol,histidine, procaine, etc.

The amount of the composition of the invention which will be effectivein the treatment, inhibition and prevention of a disease or disorderassociated with aberrant expression and/or activity of a polypeptide ofthe invention can be determined by standard clinical techniques. Inaddition, in vitro assays may optionally be employed to help identifyoptimal dosage ranges. The precise dose to be employed in theformulation will also depend on the route of administration, and theseriousness of the disease or disorder, and should be decided accordingto the judgment of the practitioner and each patient's circumstances.Effective doses may be extrapolated from dose-response curves derivedfrom in vitro or animal model test systems.

For antibodies, the dosage administered to a patient is typically 0.1mg/kg to 100 mg/kg of the patient's body weight. Preferably, the dosageadministered to a patient is between 0.1 mg/kg and 20 mg/kg of thepatient's body weight, more preferably 1 mg/kg to 10 mg/kg of thepatient's body weight. In preferred embodiments, a dose of 1, 4, 10, or20 mg/kg is administered intravenously to a patient. Generally, humanantibodies have a longer half-life within the human body than antibodiesfrom other species due to the immune response to the foreignpolypeptides. Thus, lower dosages of human antibodies and less frequentadministration is often possible. Further, the dosage and frequency ofadministration of therapeutic or pharmaceutical compositions of theinvention may be reduced by enhancing uptake and tissue penetration(e.g., into the brain) of the antibodies by modifications such as, forexample, lipidation.

The antibodies and antibody compositions of the invention may beadministered alone or in combination with other adjuvants. Adjuvantsthat may be administered with the antibody and antibody compositions ofthe invention include, but are not limited to, alum, alum plusdeoxycholate (ImmunoAg), MTP-PE (Biocine Corp.), QS21 (Genentech, Inc.),BCG, and MPL. In a specific embodiment, antibody and antibodycompositions of the invention are administered in combination with alum.In another specific embodiment, antibody and antibody compositions ofthe invention are administered in combination with QS-21. Furtheradjuvants that may be administered with the antibody and antibodycompositions of the invention include, but are not limited to,Monophosphoryl lipid immunomodulator, AdjuVax 100a, QS-21, QS-18,CRL1005, Aluminum salts, MF-59, and Virosomal adjuvant technology.Vaccines that may be administered with the antibody and antibodycompositions of the invention include, but are not limited to, vaccinesdirected toward protection against MMR (measles, mumps, rubella), polio,varicella, tetanus/diphtheria, hepatitis A, hepatitis B, haemophilusinfluenzae B, whooping cough, pneumonia, influenza, Lyme's Disease,rotavirus, cholera, yellow fever, Japanese encephalitis, poliomyelitis,rabies, typhoid fever, and pertussis, and/or PNEUMOVAX-23™. Combinationsmay be administered either concomitantly, e.g., as an admixture,separately but simultaneously or concurrently; or sequentially. Thisincludes presentations in which the combined agents are administeredtogether as a therapeutic mixture, and also procedures in which thecombined agents are administered separately but simultaneously, e.g., asthrough separate intravenous lines into the same individual.Administration “in combination” further includes the separateadministration of one of the compounds or agents given first, followedby the second.

In another specific embodiment, antibody and antibody compositions ofthe invention are used in combination with PNEUMOVAX-23™ to treat,prevent, and/or diagnose infection and/or any disease, disorder, and/orcondition associated therewith. In one embodiment, antibody and antibodycompositions of the invention are used in combination with PNEUMOVAX-23™to treat, prevent, and/or diagnose any Gram positive bacterial infectionand/or any disease, disorder, and/or condition associated therewith. Inanother embodiment, antibody and antibody compositions of the inventionare used in combination with PNEUMOVAX-23™ to treat, prevent, and/ordiagnose infection and/or any disease, disorder, and/or conditionassociated with one or more members of the genus Enterococcus and/or thegenus Streptococcus. In another embodiment, antibody and antibodycompositions of the invention are used in any combination withPNEUMOVAX-23™ to treat, prevent, and/or diagnose infection and/or anydisease, disorder, and/or condition associated with one or more membersof the Group B streptococci. In another embodiment, antibody andantibody compositions of the invention are used in combination withPNEUMOVAX-23™ to treat, prevent, and/or diagnose infection and/or anydisease, disorder, and/or condition associated with Streptococcuspneumoniae.

The antibody and antibody compositions of the invention may beadministered alone or in combination with other therapeutic agents,including but not limited to, chemotherapeutic agents, antibiotics,antivirals, steroidal and non-steroidal anti-inflammatories,conventional immunotherapeutic agents and cytokines. Combinations may beadministered either concomitantly, e.g., as an admixture, separately butsimultaneously or concurrently; or sequentially. This includespresentations in which the combined agents are administered together asa therapeutic mixture, and also procedures in which the combined agentsare administered separately but simultaneously, e.g., as throughseparate intravenous lines into the same individual. Administration “incombination” further includes the separate administration of one of thecompounds or agents given first, followed by the second.

In one embodiment, the antibody and antibody compositions of theinvention are administered in combination with other members of the TNFfamily. TNF, TNF-related or TNF-like molecules that may be administeredwith the antibody and antibody compositions of the invention include,but are not limited to, soluble forms of TNF-alpha, lymphotoxin-alpha(LT-alpha, also known as TNF-beta), LT-beta (found in complexheterotrimer LT-alpha2-beta), OPGL, FasL, CD27L, CD30L, CD40L, 4-1BBL,DcR3, OX40L, TNF-gamma (International Publication No. WO 96/14328),TRAIL (also known as AIM-1, International Publication No. WO97/33899),AIM-II (International Publication No. WO 97/34911), APRIL (J. Exp. Med.188(6):1185-1190 (1998)), endokine-alpha (International Publication No.WO 98/07880), Neutrokine-alpha (International Application PublicationNo. WO 98/18921), OPG, OX40, and nerve growth factor (NGF), and solubleforms of Fas, CD30, CD27, CD40 and 4-IBB, TR2 (International PublicationNo. WO 96/34095), DR3 (International Publication No. WO 97/33904), DR4(International Publication No. WO 98/32856), TR5 (InternationalPublication No. WO 98/30693), TR6 (International Publication No. WO98/30694), TR7 (International Publication No. WO 98/41629), TRANK, TR9(International Publication No. WO 98/56892), TR10 (InternationalPublication No. WO98/54202), 312C2 (International Publication No. WO98/06842), and TR12, and soluble forms CD154, CD70, and CD153.

In another embodiment, the compositions of the invention areadministered in combination with Neutrokine-alpha receptors and/orNeutrokine-alpha SV receptors (e.g., TACI, BCMA and BAFF-R). Inpreferred in embodiments the Neutrokine-alpha receptors and/orNeutrokine-alpha SV receptors are soluble. In other preferredembodiments the Neutrokine-alpha receptors and/or Neutrokine-alpha SVreceptors are fused to the FC region of an immunoglobulin molecule (e.g,amino acid residues 1-154 of TACI (GenBank accession number AAC51790),amino acids 1-48 of BCMA (GenBank accession number NP_(—)001183, oramino acids 1 to 81 of BAFF-R (GenBank Accession Number NP_(—)443177.)fused to the Fc region of an IgG molecule.

In a preferred embodiment, the antibody and antibody compositions of theinvention are administered in combination with CD40 ligand (CD40L), asoluble form of CD40L (e.g., AVREND™), biologically active fragments,variants, or derivatives of CD40L, anti-CD40L antibodies (e.g.,agonistic or antagonistic antibodies), and/or anti-CD40 antibodies(e.g., agonistic or antagonistic antibodies).

In an additional embodiment, the antibody and antibody compositions ofthe invention are administered alone or in combination with ananti-angiogenic agent(s). Anti-angiogenic agents that may beadministered with the antibody and antibody compositions of theinvention include, but are not limited to, Angiostatin (Entremed,Rockville, Md.), Troponin-1 (Boston Life Sciences, Boston, Mass.),anti-Invasive Factor, retinoic acid and derivatives thereof, paclitaxel(Taxol), Suramin, Tissue Inhibitor of Metalloproteinase-1, TissueInhibitor of Metalloproteinase-2, VEGI, Plasminogen ActivatorInhibitor-1, Plasminogen Activator Inhibitor-2, and various forms of thelighter “d group” transition metals.

Lighter “d group” transition metals include, for example, vanadium,molybdenum, tungsten, titanium, niobium, and tantalum species. Suchtransition metal species may form transition metal complexes. Suitablecomplexes of the above-mentioned transition metal species include oxotransition metal complexes.

Representative examples of vanadium complexes include oxo vanadiumcomplexes such as vanadate and vanadyl complexes. Suitable vanadatecomplexes include metavanadate and orthovanadate complexes such as, forexample, ammonium metavanadate, sodium metavanadate, and sodiumorthovanadate. Suitable vanadyl complexes include, for example, vanadylacetylacetonate and vanadyl sulfate including vanadyl sulfate hydratessuch as vanadyl sulfate mono- and trihydrates.

Representative examples of tungsten and molybdenum complexes alsoinclude oxo complexes. Suitable oxo tungsten complexes include tungstateand tungsten oxide complexes. Suitable tungstate complexes includeammonium tungstate, calcium tungstate, sodium tungstate dihydrate, andtungstic acid. Suitable tungsten oxides include tungsten (IV) oxide andtungsten (VI) oxide. Suitable oxo molybdenum complexes includemolybdate, molybdenum oxide, and molybdenyl complexes. Suitablemolybdate complexes include ammonium molybdate and its hydrates, sodiummolybdate and its hydrates, and potassium molybdate and its hydrates.Suitable molybdenum oxides include molybdenum (VI) oxide, molybdenum(VI) oxide, and molybdic acid. Suitable molybdenyl complexes include,for example, molybdenyl acetylacetonate. Other suitable tungsten andmolybdenum complexes include hydroxo derivatives derived from, forexample, glycerol, tartaric acid, and sugars.

A wide variety of other anti-angiogenic factors may also be utilizedwithin the context of the present invention. Representative examplesinclude, but are not limited to, platelet factor 4; protamine sulphate;sulphated chitin derivatives (prepared from queen crab shells), (Murataet al., Cancer Res. 51:22-26, 1991); Sulphated PolysaccharidePeptidoglycan Complex (SP-PG) (the function of this compound may beenhanced by the presence of steroids such as estrogen, and tamoxifencitrate); Staurosporine; modulators of matrix metabolism, including forexample, proline analogs, cishydroxyproline, d,L-3,4-dehydroproline,Thiaproline, alpha,alpha-dipyridyl, aminopropionitrile fumarate;4-propyl-5-(4-pyridinyl)-2(3H)-oxazolone; Methotrexate; Mitoxantrone;Heparin; Interferons; 2 Macroglobulin-serum; ChIMP-3 (Pavloff et al., J.Bio. Chem. 267:17321-17326, 1992); Chymostatin (Tomkinson et al.,Biochem J. 286:475-480, 1992); Cyclodextrin Tetradecasulfate;Eponemycin; Camptothecin; Fumagillin (Ingber et al., Nature 348:555-557,1990); Gold Sodium Thiomalate (“GST”; Matsubara and Ziff, J. Clin.Invest. 79:1440-1446, 1987); anticollagenase-serum; alpha2-antiplasmin(Holmes et al., J. Biol. Chem. 262(4):1659-1664, 1987); Bisantrene(National Cancer Institute); Lobenzarit disodium(N-(2)-carboxyphenyl-4-chloroanthronilic acid disodium or “CCA”;(Takeuchi et al., Agents Actions 36:312-316, 1992); andmetalloproteinase inhibitors such as BB94.

Additional anti-angiogenic factors that may also be utilized within thecontext of the present invention include Thalidomide, (Celgene, Warren,N.J.); Angiostatic steroid; AGM-1470 (H. Brem and J. Folkman J Pediatr.Surg. 28:445-51 (1993)); an integrin alpha v beta 3 antagonist (C.Storgard et al., J Clin. Invest. 103:47-54 (1999));carboxynaminolmidazole; Carboxyamidotriazole (CAI) (National CancerInstitute, Bethesda, Md.); Conbretastatin A-4 (CA4P) (OXiGENE, Boston,Mass.); Squalamine (Magainin Pharmaceuticals, Plymouth Meeting, Pa.);TNP-470, (Tap Pharmaceuticals, Deerfield, Ill.); ZD-0101 AstraZeneca(London, UK); APRA (CT2584); Benefin, Byrostatin-1 (SC339555); CGP-41251(PKC 412); CM101; Dexrazoxane (ICRF187); DMXAA; Endostatin;Flavopridiol; Genestein; GTE; ImmTher; Iressa (ZD1839); Octreotide(Somatostatin); Panretin; Penacillamine; Photopoint; PI-88; Prinomastat(AG-3340) Purlytin; Suradista (FCE26644); Tamoxifen (Nolvadex);Tazarotene; Tetrathiomolybdate; Xeloda (Capecitabine); and5-Fluorouracil.

Anti-angiogenic agents that may be administered in combination with thecompositions of the invention may work through a variety of mechanismsincluding, but not limited to, inhibiting proteolysis of theextracellular matrix, blocking the function of endothelialcell-extracellular matrix adhesion molecules, by antagonizing thefunction of angiogenesis inducers such as growth factors, and inhibitingintegrin receptors expressed on proliferating endothelial cells.Examples of anti-angiogenic inhibitors that interfere with extracellularmatrix proteolysis and which may be administered in combination with theantibody and antibody compositions of the invention include, but are notlimited to, AG-3340 (Agouron, La Jolla, Calif.), BAY-12-9566 (Bayer,West Haven, Conn.), BMS-275291 (Bristol Myers Squibb, Princeton, N.J.),CGS-27032A (Novartis, East Hanover, N.J.), Marimastat (British Biotech,Oxford, UK), and Metastat (Aeterna, St-Foy, Quebec). Examples ofanti-angiogenic inhibitors that act by blocking the function ofendothelial cell-extracellular matrix adhesion molecules and which maybe administered in combination with the antibody and antibodycompositions of the invention include, but are not limited to,EMD-121974 (Merck KcgaA Darmstadt, Germany) and Vitaxin (Ixsys, LaJolla, Calif./Medimmune, Gaithersburg, Md.). Examples of anti-angiogenicagents that act by directly antagonizing or inhibiting angiogenesisinducers and which may be administered in combination with the antibodyand antibody compositions of the invention include, but are not limitedto, Angiozyme (Ribozyme, Boulder, Colo.), Anti-VEGF antibody (Genentech,S. San Francisco, Calif.), PTK-787/ZK-225846 (Novartis, Basel,Switzerland), SU-101 (Sugen, S. San Francisco, Calif.), SU-5416(Sugen/Pharmacia Upjohn, Bridgewater, N.J.), and SU-6668 (Sugen). Otheranti-angiogenic agents act to indirectly inhibit angiogenesis. Examplesof indirect inhibitors of angiogenesis which may be administered incombination with the antibody and antibody compositions of the inventioninclude, but are not limited to, IM-862 (Cytran, Kirkland, Wash.),Interferon-alpha, IL-12 (Roche, Nutley, N.J.), and Pentosan polysulfate(Georgetown University, Washington, D.C.).

In particular embodiments, the use of antibody and antibody compositionsof the invention in combination with anti-angiogenic agents iscontemplated for the treatment, prevention, and/or amelioration of anautoimmune disease, such as for example, an autoimmune disease describedherein.

In a particular embodiment, the use of antibody and antibodycompositions of the invention in combination with anti-angiogenic agentsis contemplated for the treatment, prevention, and/or amelioration ofarthritis. In a more particular embodiment, the use of antibody andantibody compositions of the invention in combination withanti-angiogenic agents is contemplated for the treatment, prevention,and/or amelioration of rheumatoid arthritis.

In particular embodiments, the use of antibody and antibody compositionsof the invention in combination with anti-angiogenic agents iscontemplated for the treatment, prevention, and/or amelioration ofstrokes.

In another embodiment, antibody and antibody compositions of theinvention are administered in combination with an anticoagulant.Anticoagulants that may be administered with the antibody and antibodycompositions of the invention include, but are not limited to, heparin,warfarin, and aspirin. In a specific embodiment, antibody and antibodycompositions of the invention are administered in combination withheparin and/or warfarin. In another specific embodiment, antibody andantibody compositions of the invention are administered in combinationwith warfarin. In another specific embodiment, antibody and antibodycompositions of the invention are administered in combination withwarfarin and aspirin. In another specific embodiment, antibody andantibody compositions of the invention are administered in combinationwith heparin. In another specific embodiment, antibody and antibodycompositions of the invention are administered in combination withheparin and aspirin.

In another embodiment, antibody and antibody compositions of theinvention are administered in combination with an agent that suppressesthe production of anticardiolipin antibodies. In specific embodiments,the polynucleotides of the invention are administered in combinationwith an agent that blocks and/or reduces the ability of anticardiolipinantibodies to bind phospholipid-binding plasma protein beta2-glycoprotein I (b2GPI).

In certain embodiments, antibody and antibody compositions of theinvention are administered in combination with antiretroviral agents,nucleoside reverse transcriptase inhibitors, non-nucleoside reversetranscriptase inhibitors, and/or protease inhibitors. Nucleoside reversetranscriptase inhibitors that may be administered in combination withthe antibody and antibody compositions of the invention, include, butare not limited to, RETROVIR™ (zidovudine/AZT), VIDEX™ (didanosine/ddI),HIVID™ (zalcitabine/ddC), ZERIT™ (stavudine/d4T), EPIVIR™(lamivudine/3TC), and COMBIVIR™ (zidovudine/lamivudine). Non-nucleosidereverse transcriptase inhibitors that may be administered in combinationwith the antibody and antibody compositions of the invention, include,but are not limited to, VIRAMUNE™ (nevirapine), RESCRIPTOR™(delavirdine), and SUSTIVA™ (efavirenz). Protease inhibitors that may beadministered in combination with the antibody and antibody compositionsof the invention, include, but are not limited to, CRIXIVAN™(indinavir), NORVIR™ (ritonavir), INVIRASE™ (saquinavir), and VIRACEPT™(nelfinavir). In a specific embodiment, antiretroviral agents,nucleoside reverse transcriptase inhibitors, non-nucleoside reversetranscriptase inhibitors, and/or protease inhibitors may be used in anycombination with antibody and antibody compositions of the invention totreat, prevent, and/or diagnose AIDS and/or to treat, prevent, and/ordiagnose HIV infection.

In certain embodiments, antibody and antibody compositions of theinvention are administered in combination with antiretroviral agents,nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs),non-nucleoside reverse transcriptase inhibitors (NNRTIs), and/orprotease inhibitors (PIs). NRTIs that may be administered in combinationwith the compositions of the invention, include, but are not limited to,RETROVIR™ (zidovudine/AZT), VIDEX™ (didanosine/ddI), HIVID™(zalcitabine/ddC), ZERIT™ (stavudine/d4T), EPIVIR™ (lamivudine/3TC), andCOMBIVIR™ (zidovudine/lamivudine). NNRTIs that may be administered incombination with the compositions of the invention, include, but are notlimited to, VIRAMUNE™ (nevirapine), RESCRIPTOR™ (delavirdine), andSUSTIVA™ (efavirenz). Protease inhibitors that may be administered incombination with the compositions of the invention, include, but are notlimited to, CRIXIVAN™ (indinavir), NORVIR™ (ritonavir), INVIRASE™(saquinavir), and VIRACEPT™ (nelfinavir). In a specific embodiment,antiretroviral agents, nucleoside reverse transcriptase inhibitors,non-nucleoside reverse transcriptase inhibitors, and/or proteaseinhibitors may be used in any combination with compositions of theinvention to treat AIDS and/or to prevent or treat HIV infection.

Additional NRTIs include LODENOSINE™ (F-ddA; an acid-stable adenosineNRTI; Triangle/Abbott; COVIRACIL™ (emtricitabine/FTC; structurallyrelated to lamivudine (3TC) but with 3- to 10-fold greater activity invitro; Triangle/Abbott); dOTC (BCH-10652, also structurally related tolamivudine but retains activity against a substantial proportion oflamivudine-resistant isolates; Biochem Pharma); Adefovir (refusedapproval for anti-HIV therapy by FDA; Gilead Sciences); PREVEON®(Adefovir Dipivoxil, the active prodrug of adefovir; its active form isPMEA-pp); TENOFOVIR™ (bis-POC PMPA, a PMPA prodrug; Gilead); DAPD/DXG(active metabolite of DAPD; Triangle/Abbott); D-D4FC (related to 3TC,with activity against AZT/3TC-resistant virus); GW420867X (GlaxoWellcome); ZIAGEN™ (abacavir/159U89; Glaxo Wellcome Inc.); CS-87(3′azido-2′,3′-dideoxyuridine; WO 99/66936); and S-acyl-2-thioethyl(SATE)-bearing prodrug forms of b-L-FD4C and b-L-FddC (WO 98/17281).

Additional NNRTIs include COACTINON™ (Emivirine/MKC-442, potent NNRTI ofthe HEPT class; Triangle/Abbott); CAPRAVIRINE™ (AG-1549/S-1153, a nextgeneration NNRTI with activity against viruses containing the K103Nmutation; Agouron); PNU-142721 (has 20- to 50-fold greater activity thanits predecessor delavirdine and is active against K103N mutants;Pharmacia & Upjohn); DPC-961 and DPC-963 (second-generation derivativesof efavirenz, designed to be active against viruses with the K103Nmutation; DuPont); GW-420867X (has 25-fold greater activity than HBY097and is active against K103N mutants; Glaxo Wellcome); CALANOLIDE A(naturally occurring agent from the latex tree; active against virusescontaining either or both the Y181C and K103N mutations); and Propolis(WO 99/49830).

Additional protease inhibitors include LOPINAVIR™ (ABT378/r; AbbottLaboratories); BMS-232632 (an azapeptide; Bristol-Myres Squibb);TIPRANAVIR™ (PNU-140690, a non-peptic dihydropyrone; Pharmacia &Upjohn); PD-178390 (a nonpeptidic dihydropyrone; Parke-Davis); BMS232632 (an azapeptide; Bristol-Myers Squibb); L-756,423 (an indinaviranalog; Merck); DMP-450 (a cyclic urea compound; Avid & DuPont); AG-1776(a peptidomimetic with in vitro activity against proteaseinhibitor-resistant viruses; Agouron); VX-175/GW-433908 (phosphateprodrug of amprenavir; Vertex & Glaxo Welcome); CGP61755 (Ciba); andAGENERASE™ (amprenavir; Glaxo Wellcome Inc.).

Additional antiretroviral agents include fusion inhibitors/gp41 binders.Fusion inhibitors/gp41 binders include T-20 (a peptide from residues643-678 of the HIV gp41 transmembrane protein ectodomain which binds togp41 in its resting state and prevents transformation to the fusogenicstate; Trimeris) and T-1249 (a second-generation fusion inhibitor;Trimeris).

Additional antiretroviral agents include fusion inhibitors/chemokinereceptor antagonists. Fusion inhibitors/chemokine receptor antagonistsinclude CXCR4 antagonists such as AMD 3100 (a bicyclam), SDF-1 and itsanalogs, and ALX40-4C (a cationic peptide), T22 (an 18 amino acidpeptide; Trimeris) and the T22 analogs T134 and T140; CCR5 antagonistssuch as RANTES (9-68), AOP-RANTES, NNY-RANTES, and TAK-779; andCCR5/CXCR4 antagonists such as NSC 651016 (a distamycin analog). Alsoincluded are CCR2B, CCR3, and CCR6 antagonists. Chemokine receptoragonists such as RANTES, SDF-1, MIP-1a, MIP-1b, etc., may also inhibitfusion.

Additional antiretroviral agents include integrase inhibitors. Integraseinhibitors include dicaffeoylquinic (DFQA) acids; L-chicoric acid (adicaffeoyltartaric (DCTA) acid); quinalizarin (QLC) and relatedanthraquinones; ZINTEVIR™ (AR 177, an oligonucleotide that probably actsat cell surface rather than being a true integrase inhibitor; Arondex);and naphthols such as those disclosed in WO 98/50347.

Additional antiretroviral agents include hydroxyurea-like compounds suchas BCX-34 (a purine nucleoside phosphorylase inhibitor; Biocryst);ribonucleotide reductase inhibitors such as DIDOX™ (Molecules forHealth); inosine monophosphate dehydrogenase (IMPDH) inhibitors such asVX-497 (Vertex); and myvopholic acids such as CellCept (mycophenolatemofetil; Roche).

Additional antiretroviral agents include inhibitors of viral integrase,inhibitors of viral genome nuclear translocation such as arylenebis(methylketone) compounds; inhibitors of HIV entry such as AOP-RANTES,NNY-RANTES, RANTES-IgG fusion protein, soluble complexes of RANTES andglycosaminoglycans (GAG), and AMD-3100; nucleocapsid zinc fingerinhibitors such as dithiane compounds; targets of HIV Tat and Rev; andpharmacoenhancers such as ABT-378.

Other antiretroviral therapies and adjunct therapies include cytokinesand lymphokines such as MIP-1a, MIP-1b, SDF-1a, IL-2, PROLEUKIN™(aldesleukin/L2-7001; Chiron), IL-4, IL-8, IL-10, IL-12, and IL-13;interferons such as IFN-a2a; antagonists of TNFs, NFkB, GM-CSF, M-CSF,and IL-10; agents that modulate immune activation such as cyclosporinand prednisone; vaccines such as Remune™ (HIV Immunogen), APL 400-003(Apolion), recombinant gp120 and fragments, bivalent (B/E) recombinantenvelope glycoprotein, rgp120CM235, MN rgp120, SF-2 rgp120,gp120/soluble CD4 complex, Delta JR-FL protein, branched syntheticpeptide derived from discontinuous gp120 C3/C4 domain, fusion-competentimmunogens, and Gag, Pol, Nef, and Tat vaccines; gene-based therapiessuch as genetic suppressor elements (GSEs; WO 98/54366), and intrakines(genetically modified CC chemokines targeted to the ER to block surfaceexpression of newly synthesized CCR5 (Yang et al., PNAS 94:11567-72(1997); Chen et al., Nat. Med. 3:1110-16 (1997)); antibodies such as theanti-CXCR4 antibody 12G5, the anti-CCR5 antibodies 2D7, 5C7, PA8, PA9,PA10, PA11, PA12, and PA14, the anti-CD4 antibodies Q4120 and RPA-T4,the anti-CCR3 antibody 7B11, the anti-gp120 antibodies 17b, 48d,447-52D, 257-D, 268-D and 50.1, anti-Tat antibodies, anti-TNF-aantibodies, and monoclonal antibody 33A; aryl hydrocarbon (AH) receptoragonists and antagonists such as TCDD, 3,3′,4,4′,5-pentachlorobiphenyl,3,3′,4,4′-tetrachlorobiphenyl, and a-naphthoflavone (WO 98/30213); andantioxidants such as g-L-glutamyl-L-cysteine ethyl ester (g-GCE; WO99/56764).

In other embodiments, antibody and antibody compositions of theinvention may be administered in combination with anti-opportunisticinfection agents. Anti-opportunistic agents that may be administered incombination with the antibody and antibody compositions of theinvention, include, but are not limited to,TRIMETHOPRIM-SULFAMETHOXAZOLE™, DAPSONE™, PENTAMIDINE™, ATOVAQUONE™,ISONIAZID™, RIFAMPIN™, PYRAZINAMIDE™, ETHAMBUTOL™, RIFABUTIN™,CLARITHROMYCIN™, AZITHROMYCIN™, GANCICLOVIR™, FOSCARNET™, CIDOFOVIR™,FLUCONAZOLE™, ITRACONAZOLE™, KETOCONAZOLE™, ACYCLOVIR™, FAMCICOLVIR™,PYRIMETHAMINE™, LEUCOVORIN™, NEUPOGEN™ (filgrastim/G-CSF), and LEUKINE™(sargramostim/GM-CSF). In a specific embodiment, antibody and antibodycompositions of the invention are used in any combination withTRIMETHOPRIM-SULFAMETHOXAZOLE™, DAPSONE™, PENTAMIDINE™, and/orATOVAQUONE™ to prophylactically treat, prevent, and/or diagnose anopportunistic Pneumocystis carinii pneumonia infection. In anotherspecific embodiment, antibody and antibody compositions of the inventionare used in any combination with ISONIAZID™, RIFAMPIN™, PYRAZINAMIDE™,and/or ETHAMBUTOL™ to prophylactically treat, prevent, and/or diagnosean opportunistic Mycobacterium avium complex infection. In anotherspecific embodiment, antibody and antibody compositions of the inventionare used in any combination with RIFABUTIN™, CLARITHROMYCIN™, and/orAZITHROMYCIN™ to prophylactically treat, prevent, and/or diagnose anopportunistic Mycobacterium tuberculosis infection. In another specificembodiment, antibody and antibody compositions of the invention are usedin any combination with GANCICLOVIR™, FOSCARNET™, and/or CIDOFOVIR™ toprophylactically treat, prevent, and/or diagnose an opportunisticcytomegalovirus infection. In another specific embodiment, antibody andantibody compositions of the invention are used in any combination withFLUCONAZOLE™, ITRACONAZOLE™, and/or KETOCONAZOLE™ to prophylacticallytreat, prevent, and/or diagnose an opportunistic fungal infection. Inanother specific embodiment, antibody and antibody compositions of theinvention are used in any combination with ACYCLOVIR™ and/orFAMCICOLVIR™ to prophylactically treat, prevent, and/or diagnose anopportunistic herpes simplex virus type I and/or type II infection. Inanother specific embodiment, antibody and antibody compositions of theinvention are used in any combination with PYRIMETHAMINE™ and/orLEUCOVORIN™ to prophylactically treat, prevent, and/or diagnose anopportunistic Toxoplasma gondii infection. In another specificembodiment, antibody and antibody compositions of the invention are usedin any combination with LEUCOVORIN™ and/or NEUPOGEN™ to prophylacticallytreat, prevent, and/or diagnose an opportunistic bacterial infection.

In a further embodiment, the antibody and antibody compositions of theinvention are administered in combination with an antiviral agent.Antiviral agents that may be administered with the antibody and antibodycompositions of the invention include, but are not limited to,acyclovir, ribavirin, amantadine, and remantidine.

In a further embodiment, the antibody and antibody compositions of theinvention are administered in combination with an antibiotic agent.Antibiotic agents that may be administered with the antibody andantibody compositions of the invention include, but are not limited to,amoxicillin, aminoglycosides, beta-lactam (glycopeptide),beta-lactamases, Clindamycin, chloramphenicol, cephalosporins,ciprofloxacin, ciprofloxacin, erythromycin, fluoroquinolones,macrolides, metronidazole, penicillins, quinolones, rifampin,streptomycin, sulfonamide, tetracyclines, trimethoprim,trimethoprim-sulfamthoxazole, and vancomycin.

Conventional nonspecific immunosuppressive agents, that may beadministered in combination with the antibody and antibody compositionsof the invention include, but are not limited to, steroids,cyclosporine, cyclosporine analogs cyclophosphamide, cyclophosphamideIV, methylprednisolone, prednisolone, azathioprine, FK-506,15-deoxyspergualin, and other immunosuppressive agents that act bysuppressing the function of responding T cells. Other immunosuppressiveagents, that may be administered in combination with the compositions ofthe invention include, but are not limited to, prednisolone,methotrexate, thalidomide, methoxsalen, rapamycin, leflunomide,mizoribine (BREDININ™), brequinar, deoxyspergualin, and azaspirane (SKF105685).

In specific embodiments, antibody and antibody compositions of theinvention are administered in combination with immunosuppressantsImmunosuppressants preparations that may be administered with theantibody and antibody compositions of the invention include, but are notlimited to, ORTHOCLONE™ (OKT® 3 (muromonab-CD3), SANDIMMUNE™, NEORAL™,SANGDYA™ (cyclosporine), PROGRAF® (FK506, tacrolimus), CELLCEPT®(mycophenolate motefil, of which the active metabolite is mycophenolicacid), IMURAN™ (azathioprine), glucorticosteroids, adrenocorticalsteroids such as DELTASONE™ (prednisone) and HYDELTRASOL™(prednisolone), FOLEX™ and MEXATE™ (methotrxate), OXSORALEN-ULTRA™(methoxsalen) and RAPAMUNE™ (sirolimus). In a specific embodiment,immunosuppressants may be used to prevent rejection of organ or bonemarrow transplantation.

In a preferred embodiment, the antibody and antibody compositions of theinvention are administered in combination with steroid therapy. Steroidsthat may be administered in combination with the antibody and antibodycompositions of the invention, include, but are not limited to, oralcorticosteroids, prednisone, and methylprednisolone (e.g., IVmethylprednisolone). In a specific embodiment, antibody and antibodycompositions of the invention are administered in combination withprednisone. In a further specific embodiment, the antibody and antibodycompositions of the invention are administered in combination withprednisone and an immunosuppressive agent Immunosuppressive agents thatmay be administered with the antibody and antibody compositions of theinvention and prednisone are those described herein, and include, butare not limited to, azathioprine, cylophosphamide, and cyclophosphamideIV. In a another specific embodiment, antibody and antibody compositionsof the invention are administered in combination withmethylprednisolone. In a further specific embodiment, the antibody andantibody compositions of the invention are administered in combinationwith methylprednisolone and an immunosuppressive agent Immunosuppressiveagents that may be administered with the antibody and antibodycompositions of the invention and methylprednisolone are those describedherein, and include, but are not limited to, azathioprine,cylophosphamide, and cyclophosphamide IV.

In a preferred embodiment, the antibody and antibody compositions of theinvention are administered in combination with an antimalarial.Antimalarials that may be administered with the antibody and antibodycompositions of the invention include, but are not limited to,hydroxychloroquine, chloroquine, and/or quinacrine.

In a preferred embodiment, the antibody and antibody compositions of theinvention are administered in combination with an NSAID.

In a nonexclusive embodiment, the antibody and antibody compositions ofthe invention are administered in combination with one, two, three,four, five, ten, or more of the following drugs: NRD-101 (Hoechst MarionRoussel), diclofenac (Dimethaid), oxaprozin potassium (Monsanto),mecasermin (Chiron), T-614 (Toyama), pemetrexed disodium (Eli Lilly),atreleuton (Abbott), valdecoxib (Monsanto), eltenac (Byk Gulden),campath, AGM-1470 (Takeda), CDP-571 (Celltech Chiroscience), CM-101(CarboMed), ML-3000 (Merckle), CB-2431 (KS Biomedix), CBF-BS2 (KSBiomedix), IL-1Ra gene therapy (Valentis), JTE-522 (Japan Tobacco),paclitaxel (Angiotech), DW-166HC (Dong Wha), darbufelone mesylate(Warner-Lambert), soluble TNF receptor 1 (synergen; Amgen), IPR-6001(Institute for Pharmaceutical Research), trocade (Hoffman-La Roche),EF-5 (Scotia Pharmaceuticals), BIIL-284 (Boehringer Ingelheim),BIIF-1149 (Boehringer Ingelheim), LeukoVax (Inflammatics), MK-663(Merck), ST-1482 (Sigma-Tau), and butixocort propionate (WarnerLambert).

In one embodiment, the compositions of the invention are administered incombination with one or more of the following drugs: Infliximab (alsoknown as Remicade™ Centocor, Inc.), Trocade (Roche, RO-32-3555),Leflunomide (also known as Arava™ from Hoechst Marion Roussel), Kineret™(an IL-1 Receptor antagonist also known as Anakinra from Amgen, Inc.),SCIO-469 (p38 kinase inhibitor from Scios, Inc), Humira® (adalimumabfrom Abbott Laboratories) and/or ASLERA™ (prasterone,dehydroepiandrosterone, GL701) from Genelabs Technologies Inc.

In a preferred embodiment, the antibody and antibody compositions of theinvention are administered in combination with one, two, three, four,five or more of the following drugs: methotrexate, sulfasalazine, sodiumaurothiomalate, auranofin, cyclosporine, penicillamine, azathioprine, anantimalarial drug (e.g., as described herein), cyclophosphamide,chlorambucil, gold, ENBREL™ (Etanercept), anti-TNF antibody, LJP 394 (LaJolla Pharmaceutical Company, San Diego, Calif.) and prednisolone.

In a more preferred embodiment, the antibody and antibody compositionsof the invention are administered in combination with an antimalarial,methotrexate, anti-TNF antibody, ENBREL™ and/or suflasalazine. In oneembodiment, the antibody and antibody compositions of the invention areadministered in combination with methotrexate. In another embodiment,the antibody and antibody compositions of the invention are administeredin combination with anti-TNF antibody. In another embodiment, theantibody and antibody compositions of the invention are administered incombination with methotrexate and anti-TNF antibody. In anotherembodiment, the antibody and antibody compositions of the invention areadministered in combination with suflasalazine. In another specificembodiment, the antibody and antibody compositions of the invention areadministered in combination with methotrexate, anti-TNF antibody, andsuflasalazine. In another embodiment, the antibody and antibodycompositions of the invention are administered in combination ENBREL™.In another embodiment, the antibody and antibody compositions of theinvention are administered in combination with ENBREL™ and methotrexate.In another embodiment, the antibody and antibody compositions of theinvention are administered in combination with ENBREL™, methotrexate andsuflasalazine. In another embodiment, the antibody and antibodycompositions of the invention are administered in combination withENBREL™, methotrexate and suflasalazine. In other embodiments, one ormore antimalarials is combined with one of the above-recitedcombinations. In a specfic embodiment, the antibody and antibodycompositions of the invention are administered in combination with anantimalarial (e.g., hydroxychloroquine), ENBREL™, methotrexate andsuflasalazine. In another specfic embodiment, the antibody and antibodycompositions of the invention are administered in combination with anantimalarial (e.g., hydroxychloroquine), sulfasalazine, anti-TNFantibody, and methotrexate.

In an additional embodiment, antibody and antibody compositions of theinvention are administered alone or in combination with one or moreintravenous immune globulin preparations. Intravenous immune globulinpreparations that may be administered with the antibody and antibodycompositions of the invention include, but not limited to, GAMMAR™,IVEEGAM™, SANDOGLOBULIN™, GAMMAGARD S/D™, and GAMIMUNE™. In a specificembodiment, antibody and antibody compositions of the invention areadministered in combination with intravenous immune globulinpreparations in transplantation therapy (e.g., bone marrow transplant).

In an additional embodiment, the antibody and antibody compositions ofthe invention are administered alone or in combination with ananti-inflammatory agent. Anti-inflammatory agents that may beadministered with the antibody and antibody compositions of theinvention include, but are not limited to, glucocorticoids and thenonsteroidal anti-inflammatories, aminoarylcarboxylic acid derivatives,arylacetic acid derivatives, arylbutyric acid derivatives,arylcarboxylic acids, arylpropionic acid derivatives, pyrazoles,pyrazolones, salicylic acid derivatives, thiazinecarboxamides,e-acetamidocaproic acid, S-adenosylmethionine, 3-amino-4-hydroxybutyricacid, amixetrine, bendazac, benzydamine, bucolome, difenpiramide,ditazol, emorfazone, guaiazulene, nabumetone, nimesulide, orgotein,oxaceprol, paranyline, perisoxal, pifoxime, proquazone, proxazole, andtenidap.

In specific embodiments, the compositions of the invention areadministered alone or in combination with anti-CD4 antibody. In oneembodiment, coadministration of the compositions of the invention withanti-CD4 antibody is envisoned for treatment of rheumatoid arthritis.

In specific embodiments, the compositions of the invention areadministered alone or in combination with anti-IL-15 antibody. In oneembodiment, coadministration of the compositions of the invention withanti-IL-15 antibody is envisoned for treatment of rheumatoid arthritis.

In specific embodiments, the compositions of the invention areadministered alone or in combination with CTLA4-Ig and LEA29Y. In oneembodiment, coadministration of the compositions of the invention withCTLA4-Ig and LEA29Y is envisoned for treatment of rheumatoid arthritis.

In specific embodiments, the compositions of the invention areadministered alone or in combination with anti-IL-6 Receptor antibody.In one embodiment, coadministration of the compositions of the inventionwith anti-IL-6 Receptor antibody is envisoned for treatment ofrheumatoid arthritis.

In specific embodiments, the compositions of the invention areadministered alone or in combination with anti-C5 (complement component)antibody. In one embodiment, coadministration of the compositions of theinvention with anti-C5 antibody is envisoned for treatment of rheumatoidarthritis.

In specific embodiments, the compositions of the invention areadministered alone or in combination with complement cascade inhibitors.Complement cascade inhibitors include, but are notl imited to,anti-properdin antibodies (Gliatech); TP-10, a recombinant soluble typeI complement receptor (AVANT Immunotheragenetics Inc.); Pexelizmab, aComplement C5 inhibitor (Alexion Pharmaceuticals Inc.); and 5G1.1, amonoclonal antibody that prevents cleavage of complement component C5into its pro-inflammatory components. In one embodiment,coadministration of the compositions of the invention with complementcascade inhibitors are is envisoned for treatment of Inflammation,Rheumatoid arthritis, and/or cardiovascular disorders.

In another embodiment, compostions of the invention are administered incombination with a chemotherapeutic agent. Chemotherapeutic agents thatmay be administered with the antibody and antibody compositions of theinvention include, but are not limited to, antibiotic derivatives (e.g.,doxorubicin, bleomycin, daunorubicin, and dactinomycin); antiestrogens(e.g., tamoxifen); antimetabolites (e.g., fluorouracil, 5-FU,methotrexate, floxuridine, interferon alpha-2b, glutamic acid,plicamycin, mercaptopurine, and 6-thioguanine); cytotoxic agents (e.g.,carmustine, BCNU, lomustine, CCNU, cytosine arabinoside,cyclophosphamide, estramustine, hydroxyurea, procarbazine, mitomycin,busulfan, cis-platin, and vincristine sulfate); hormones (e.g.,medroxyprogesterone, estramustine phosphate sodium, ethinyl estradiol,estradiol, megestrol acetate, methyltestosterone, diethylstilbestroldiphosphate, chlorotrianisene, and testolactone); nitrogen mustardderivatives (e.g., mephalen, chorambucil, mechlorethamine (nitrogenmustard) and thiotepa); steroids and combinations (e.g., bethamethasonesodium phosphate); and others (e.g., dicarbazine, asparaginase,mitotane, vincristine sulfate, vinblastine sulfate, and etoposide).

In a specific embodiment, compositions of the invention are administeredin combination with CHOP (cyclophosphamide, doxorubicin, vincristine,and prednisone) or combination of one or more of the components of CHOP.In one embodiment, the compositions of the invention are administered incombination with anti-CD20 antibodies, human monoclonal anti-CD20antibodies. In another embodiment, the compositions of the invention areadministered in combination with anti-CD20 antibodies and CHOP, oranti-CD20 antibodies and any combination of one or more of thecomponents of CHOP, particularly cyclophosphamide and/or prednisone. Ina specific embodiment, compositions of the invention are administered incombination with Rituximab. In a further embodiment, compositions of theinvention are administered with Rituximab and CHOP, or Rituximab and anycombination of one or more of the components of CHOP, particularlycyclophosphamide and/or prednisone. In a specific embodiment,compositions of the invention are administered in combination withtositumomab (anti-CD20 antibody from Coulter Pharmaceuticals, SanFrancisco, Calif.). In a further embodiment, compositions of theinvention are administered with tositumomab and CHOP, or tositumomab andany combination of one or more of the components of CHOP, particularlycyclophosphamide and/or prednisone. Tositumomab may optionally beassociated with 131I. The anti-CD20 antibodies may optionally beassociated with radioisotopes, toxins or cytotoxic prodrugs.

In another specific embodiment, the compositions of the invention areadministered in combination Zevalin™. In a further embodiment,compositions of the invention are administered with Zevalin™ and CHOP,or Zevalin™ and any combination of one or more of the components ofCHOP, particularly cyclophosphamide and/or prednisone. Zevalin™ may beassociated with one or more radisotopes. Particularly preferred isotopesare 90Y and 111In.

In additional preferred embodiments, compositions of the invention areadministered in combination with Rituximab (Rituxan™) and/or IbritumomabTiuxetan (Zevalin™, e.g., either (In-111) Ibritumomab Tiuxetan or (Y-90)Ibritumomab Tiuxetan). In a specific embodiment, compositions of theinvention are administered in combination with Rituximab and/orIbritumomab Tiuxetan for the treatment of non-Hodgkin's lymphoma.

In additional preferred embodiments, compositions of the invention areadministered in combination with imatinib mesylate (Gleevec®:4-[(4-Methyl-1-piperazinyl)methyl]-N-[4-methyl-3-[[4-(3-pyridinyl)-2-pyrimidinyl]amino]-phenyl]benzamidemethanesulfonate). In a specific embodiment, compositions of theinvention are administered in combination with imatinib mesylate for thetreatment of chronic myelogenous leukemia.

In additional preferred embodiments, compositions of the invention areadministered in combination with bortezomib (Velcade™[(1R)-3-methyl-1-[[(2S)-1-oxo-3-phenyl-2-[(pyrazinylcarbonyl)amino]propyl]amino]butyl]boronicacid). In a specific embodiment, compositions of the invention areadministered in combination with bortezomib for the treatment ofmultiple myeloma.

In additional preferred embodiments, compositions of the invention areadministered in combination with Alemtuzumab (Campath®). In a specificembodiment, compositions of the invention are administered incombination with Alemtuzumab for the treatment of chronic lymphocyticleukemia.

In additional preferred embodiments, compositions of the invention areadministered in combination with fludarabine phosphate (Fludara®:9H-Purin-6-amine, 2-fluoro-9-(5-O-phosphono-b-D-arabinofuranosyl)(2-fluoro-ara-AMP)). In a specific embodiment, compositions of theinvention are administered in combination with fludarabine phosphate forthe treatment of chronic lymphocytic leukemia.

In further particular embodiments, compositions of the present inventionare used to treat, ameliorate and/or prevent hematological cancers.Compositions of the present invention may be used in combination withone or more surgical and/or radiological procedures and/or therapeuticagents to treat, ameliorate and/or prevent hematological cancers.Hematological cancers which may be treated using compositions of thepresent invention include, but are not limited to, non-Hodgkin'slymphoma (e.g., small lymphocytic lymphoma, follicular center celllymphoma, lymphoplasmacytoid lymphoma, marginal zone lymphoma, mantlecell lymphoma, immunoblastic lymphoma, burkitt's lymphoma, lymphoblasticlymphoma, peripheral T-cell lymphoma, anaplastic large cell lymphoma andintestinal T-cell lymphoma), leukemia, acute lymphocytic leukemia,chronic lymphocytic leukemia and plasma cell neoplasms includingmultiple myeloma.

In preferred embodiments, compositions of the present invention are usedto treat, ameliorate and/or prevent hematological cancers. compositionsof the present invention may be used in combination with one or moresurgical and/or radiological procedures and/or therapeutic agents totreat, ameliorate and/or prevent hematological cancers. Hematologicalcancers which may be treated using compositions of the present inventioninclude, but are not limited to, non-Hodgkin's lymphoma (e.g., smalllymphocytic lymphoma, follicular center cell lymphoma,lymphoplasmacytoid lymphoma, marginal zone lymphoma, mantle celllymphoma, immunoblastic lymphoma, burkitt's lymphoma, lymphoblasticlymphoma, peripheral T-cell lymphoma, anaplastic large cell lymphoma andintestinal T-cell lymphoma), leukemia, acute lymphocytic leukemia,chronic lymphocytic leukemia and plasma cell neoplasms includingmultiple myeloma.

In one preferred embodiment, compositions of the present invention areused to treat plasma cell neoplasms. In a specific embodiment, thatplasma cell neoplasm is multiple myeloma.

In another preferred embodiment, compositions of the present inventionare used to treat non-Hodgkin's lymphoma.

In another preferred embodiment, compositions of the present inventionare used to treat leukemia. In a specific embodiment, that leukemia isacute lymphocytic leukemia. In another specific embodiment, thatleukemia is chronic lymphocytic leukemia.

Compositions of the present invention may be administered in combinationwith one or more surgical and/or radiological procedures useful in thetreatment of hematological cancer including, but not limited to, bonemarrow transplantation, external beam radiation and total bodyirradiation.

In preferred embodiments, compositions of the present invention areadministered in combination with one or more surgical and/orradiological procedures useful in the treatment of hematological cancerincluding, but not limited to, bone marrow transplantation, externalbeam radiation and total body irradiation.

In one preferred embodiment, compositions of the present invention maybe administered in combination with one or more surgical and/orradiological procedures useful in the treatment of multiple myelomaincluding, but not limited to, allogeneic bone marrow transplantationand peripheral stem cell support.

In another preferred embodiment, compositions of the present inventionmay be administered in combination with one or more surgical and/orradiological procedures useful in the treatment of non-Hodgkin'slymphoma including, but not limited to, allogeneic bone marrowtransplantation and peripheral stem cell support.

In further specific embodiments, compositions of the present inventionmay be administered in combination with one or more surgical and/orradiological procedures useful in the treatment of leukemia including,but not limited to, allogeneic bone marrow transplantation andperipheral stem cell support. In one specific preferred embodiment,compositions of the present invention of the invention are used to treatacute lymphocytic leukemia (ALL). In another specific preferredembodiment, compositions of the present invention are used to treatchronic lymphocytic leukemia (CLL).

Compositions of the present invention may be administered in combinationwith one or more therapeutic agents useful in the treatment of multiplemyeloma including, but not limited to, Alkylating agents,Anthracyclines, Carmustine (DTI-015, BCNU, BiCNU, Gliadel Wafer®),Cyclophosphamide (Cytoxan®, Neosar®, CTX), Dexamethasone (Decadron®),Doxorubicin (Adriamycin®, Doxil®, Rubex®), Melphalan (L-PAM, Alkeran®,Phenylalanine mustard), Prednisone, Thalidomide and Vincristine(Oncovorin®, Onco TCS®, VCR, Leurocristine®).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agents in the treatment, amelioration and/or prevention ofmultiple myeloma.

Preferred combinations of therapeutic agents useful in the treatment ofmultiple myeloma which may be administered in combination withcompositions of the present invention include, but are not limited to,Cyclophosphamide+Prednisone, Melphalan+Prednisone (MP),Vincristine+Adriamycin®+Dexamethasone (VAD),Vincristine+Carmustine+Melphalan+Cyclophosphamide+Prednisone (VBMCP; theM2 protocol), and Vincristine+Melphalan+Cyclophosphamide+Prednisonealternating with Vincristine+Carmustine+Doxorubicin+Prednisone(VMCP/VBAP).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agent combinations in the treatment, amelioration and/orprevention of multiple myeloma.

Compositions of the present invention may be administered in combinationwith one or more therapeutic agents useful in the treatment ofnon-Hodgkin's lymphoma including, but not limited to,2-chlorodeoxyadenosine, Amifostine (Ethyol®, Ethiofos®, WR-272),Bexarotene (Targretin®, Targretin gel®, Targretin oral®, LGD1069),Bleomycin (Blenoxane®), Busulfan (Busulfex®, Myleran®), Carboplatin(Paraplatin®, CBDCA), Carmustine (DTI-015, BCNU, BiCNU, Gliadel Wafer®),Chlorambucil (Leukeran®), Cisplatin (Platinol®, CDDP), Cladribine(2-CdA, Leustatin®), Cyclophosphamide (Cytoxan®, Neosar®, CTX),Cytarabine (Cytosar-U®, ara-C, cytosine arabinoside, DepoCyt®),Dacarbazine (DTIC), Daunorubicin (Daunomycin, DaunoXome®, Daunorubicin®,Cerubidine®), Denileukin diftitox (Ontak®), Dexamethasone (Decadron®),Dolasetron mesylate (Anzemet®), Doxorubicin (Adriamycin®, Doxil®,Rubex®), Erythropoietin (EPO®, Epogen®, Procrit®), Etoposide phosphate(Etopophos®), Etoposide (VP-16, Vepesid®), Fludarabine (Fludara®, FAMP),Granisetron (Kytril®), Hydrocortisone, Idarubicin (Idamycin®, DMDR,IDA), Ifosfamide (IFEX®), Interferon alpha (Alfaferone®, Alpha-IF®),Interferon alpha 2a (Intron A®), Mechlorethamine (Nitrogen Mustard, HN2,Mustargen®), Melphalan (L-PAM, Alkeran®, Phenylalanine mustard),Methotrexate® (MTX, Mexate®, Folex®), Methylprednisolone (Solumedrol®),Mitoxantrone (Novantrone®, DHAD), Ondansetron (Zofran®), Pentostatin(Nipent®, 2-deoxycoformycin), Perfosfamide(4-hydroperoxycyclophosphamide, 4-HC), Prednisone, Procarbazine(Matulane®), Rituximab® (Rituxan®, anti-CD20 MAb), Thiotepa(triethylenethiophosphaoramide, Thioplex®), Topotecan (Hycamtin®,SK&F-104864, NSC-609699, Evotopin®), Vinblastine (Velban®, VLB),Vincristine (Oncovin®, Onco TCS®, VCR, Leurocristine®) and Vindesine(Eldisine®, Fildesin®).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agents in the treatment, amelioration and/or prevention ofnon-Hodgkin's lymphoma.

Preferred combinations of therapeutic agents useful in the treatment ofnon-Hodgkin's lymphoma which may be administered in combination withcompositions of the present invention include, but are not limited to,Adriamycin®+Blenoxane+Vinblastine+Dacarbazine (ABVD), Anti-idiotypetherapy (BsAb)+Interferon alpha, Anti-idiotype therapy(BsAb)+Chlorambucil, Anti-idiotype therapy (BsAb)+Interleukin-2, BCNU(Carmustine)+Etoposide+Ara-C (Cytarabine)+Melphalen (BEAM),Bleomycin+Etoposide+Adriamycin+Cyclophosphamide+Vincristine+Procarbazine+Prednisone(BEACOPP), Bryostatin+Vincristine, Cyclophosphamide+BCNU(Carmustine)+VP-16 (Etoposide) (CBV),Cyclophosphamide+Vincristine+Prednisone (CVP),Cyclophosphamide+Adriamycin® (Hydroxyldaunomycin)+Vincristine(Oncovorin)+Prednisone (CHOP), Cyclophosphamide+Novantrone®(Mitoxantrone)+Vincristine (Oncovorin)+Prednisone (CNOP),Cyclophosphamide+Doxorubicin+Teniposide+Prednisone,Cyclophosphamide+Adriamycin® (Hydroxyldaunomycin)+Vincristine(Oncovorin)+Prednisone+Rituximab (CHOP+Rituximab),Cyclophosphamide+Doxorubicin+Teniposide+Prednisone+Interferon alpha,Cytarabine+Bleomycin+Vincristine+Methotrexate (CytaBOM),Dexamethasone+Cytarabine+Cisplatin (DHAP),Dexamethasone+Ifosfamide+Cisplatin+Etoposide (DICE),Doxorubicin+Vinblastine+Mechlorethamine+Vincristine+Bleomycin+Etoposide+Prednisone(Stanford V), Etoposide+Vinblastine+Adriamycin (EVA),Etoposide+Methylprednisone+Cytarabine+Cisplatin (ESHAP),Etoposide+Prednisone+Ifosfamide+Cisplatin (EPIC), Fludarabine,Mitoxantrone+Dexamethasone (FMD), Fludarabine, Dexamethasone, Cytarabine(ara-C), +Cisplatin (Platinol®) (FluDAP), Ifosfamide+Cisplatin+Etoposide(ICE), Mechlorethamine+Oncovin® (Vincristine)+Procarbazine+Prednisone(MOPP), Mesna+Ifosfamide+Idarubicin+Etoposide (MIZE), Methotrexate withleucovorinrescue+Bleomycin+Adriamycin+Cyclophosphamide+Oncovorin+Dexamethasone(m-BACOD), Prednisone+Methotrexate+Adriamycin+Cyclophosphamide+Etoposide(ProMACE), Thiotepa+Busulfan+Cyclophosphamide,Thiotepa+Busulfan+Melphalan, Topotecan+Paclitaxel, and Vincristine(Oncovin®)+Adriamycin®+Dexamethasone (VAD).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agent combinations in the treatment, amelioration and/orprevention of non-Hodgkin's lymphoma.

Further examples of therapeutic agents useful in the treatment ofnon-Hodgkin's lymphoma which may be administered in combination withcompositions of the present invention include, but are not limited to,A007 (4-4′-dihydroxybenzophenone-2,4-dinitrophenylhydrazone), AG-2034(AG-2024, AG-2032, GARFT [glycinamide ribonucleoside transformylase]inhibitor), Aldesleukin (IL-2, Proleukin®), Alemtuzumab (Campath®),Alitretinoin (Panretin®, LGN-1057), Altretamine (Hexalen®,hexamethylmelamine, Hexastat®), Aminocamptothecin (9-AC,9-Aminocamptothecin, NSC 603071), Anti-CD19/CD3 MAb (anti-CD19/CD3 scFv,anti-NHL MAb), Anti-idiotype therapy (BsAb), Arabinosylguanine (Ara-G,GW506U78), Arsenic trioxide (Trisenox®, ATO), B43-Genistein (anti-CD19Ab/genistein conjugate), B7 antibody conjugates, Betathine (Beta-LT), BLymphocyte Stimulator antagonists, Bryostatin-1 (Bryostatin®, BMY-45618,NSC-339555), CHML (Cytotropic Heterogeneous Molecular Lipids),Clofarabine (chloro-fluoro-araA), Daclizumab (Zenapax®), Depsipeptide(FR901228, FK228), Dolastatin-10 (DOLA-10, NSC-376128), Epirubicin(Ellence®, EPI, 4′ epi-doxorubicin), Epratuzumab (Lymphocide®, humanizedanti-CD22, HAT), Fly3/flk2 ligand (Mobista®), G3139 (Genasense®,GentaAnticode®, Bcl-2 antisense), Hu1D10 (anti-HLA-DR MAb, SMART 1D10),HumaLYM (anti-CD20 MAb), Ibritumomab tiuxetan (Zevalin®), Interferongamma (Gamma-interferon, Gamma 100®, Gamma-IF), Irinotecan (Camptosar®,CPT-11, Topotecin®, CaptoCPT-1), ISIS-2053, ISIS-3521 (PKC-alphaantisense), Lmb-2 immunotoxin (anti-CD25 recombinant immuno toxin,anti-Tac(Fv)-PE38), Leuvectin® (cytofectin+IL-2 gene, IL-2 genetherapy), Lym-1 (131-I LYM-1), Lymphoma vaccine (Genitope), Nelarabine(Compound 506, U78), Neugene compounds (Oncomyc-NG®, Resten-NG®, mycantisense), NovoMAb-G2 scFv (NovoMAb-G2 IgM), O6-benzylguanine (BG,Procept®), Oxaliplatin (Eloxatine®, Eloxatin®), Paclitaxel (Paxene®,Taxol®), Paclitaxel-DHA (Taxoprexin®), Peldesine (BCX-34, PNPinhibitor), Rebeccamycin and Rebeccamycin analogues, SCH-66336,Sobuzoxane (MST-16, Perazolin®), SU5416 (Semaxanib®, VEGF inhibitor),TER-286, Thalidomide, TNP-470 (AGM-1470), Tositumomab (Bexxar®),Valspodar (PSC 833), Vaxid (B-cell lymphoma DNA vaccine), Vinorelbine(Navelbine®), WF10 (macrophage regulator) and XR-9576 (XR-9351,P-glycoprotein/MDR inhibitor).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agents in the treatment, amelioration and/or prevention ofnon-Hodgkin's lymphoma.

Compositions of the present invention may be administered in combinationwith one or more therapeutic agents useful in the treatment of acutelymphocytic leukemia including, but not limited to, Amsacrine,Carboplatin (Paraplatin®, CBDCA), Carmustine (DTI-015, BCNU, BiCNU,Gliadel Wafer®), Cholecaliferol, Cyclophosphamide (Cytoxan®, Neosar®,CTX), Cytarabine (Cytosar-U®, ara-C, cytosine arabinoside, DepoCyt®),Daunorubicin (Daunomycin, DaunoXome®, Daunorubicin®, Cerubidine®),Dexamethasone (Decadron®), Doxorubicin (Adriamycin®, Doxil®, Rubex®),Etoposide (VP-16, Vepesid®), Filgrastam® (Neupogen®, G-CSF, Leukine®),Fludarabine (Fludara®, FAMP), Idarubicin (Idamycin®, DMDR, IDA),Ifosfamide (IFEX®), Imatinib mesylate (STI-571, Imatinib®, Glivec®,Gleevec®, Abl tyrosine kinase inhibitor), Interferon gamma(Gamma-interferon, Gamma 100®, Gamma-IF), L-asparaginase (Elspar®,Crastinin®, Asparaginase medac®, Kidrolase®), Mercaptopurine(6-mercaptopurine, 6-MP), Methotrexate® (MTX, Mexate®, Folex®),Mitoxantrone (Novantrone®, DHAD), Pegaspargase® (Oncospar®), Prednisone,Retinoic acid, Teniposide (VM-26, Vumon®), Thioguanine (6-thioguanine,6-TG), Topotecan (Hycamtin®, SK&F-104864, NSC-609699, Evotopin®),Tretinoin (Retin-A®, Atragen®, ATRA, Vesanoid®) and Vincristine(Oncovorin®, Onco TCS®, VCR, Leurocristine®).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agents in the treatment, amelioration and/or prevention ofacute lymphocytic leukemia.

Further examples of therapeutic agents useful in the treatment of acutelymphocytic leukemia which may be administered in combination withcompositions of the present invention include, but are not limited to,Aminocamptothecin (9-AC, 9-Aminocamptothecin, NSC 603071), Aminopterin,Annamycin (AR-522, annamycin LF, Aronex®), Arabinosylguanine (Ara-G,GW506U78, Nelzarabine®), Arsenic trioxide (Trisenox®, ATO, Atrivex®),B43-Genistein (anti-CD19 Ab/genistein conjugate), B43-PAP (anti-CD19Ab/pokeweed antiviral protein conjugate), Cordycepin, CS-682, Decitabine(5-aza-2′-deoxyytidine), Dolastatin-10 (DOLA-10, NSC-376128), G3139(Genasense®, GentaAnticode®, Bcl-2 antisense), Irofulven (MGI-114,Ivofulvan, Acylfulvene analogue), MS-209, Phenylbutyrate, Quinine,TNP-470 (AGM-1470, Fumagillin), Trimetrexate (Neutrexin®), Troxacitabine(BCH-204, BCH-4556, Troxatyl®), UCN-01 (7-hydroxystaurosporine),WHI-P131 and WT1 Vaccine.

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agents in the treatment, amelioration and/or prevention ofacute lymphocytic leukemia.

Preferred combinations of therapeutic agents useful in the treatment ofacute lymphocytic leukemia which may be administered in combination withcompositions of the present invention include, but are not limited to,Carboplatin+Mitoxantrone, Carmustine+Cyclophosphamide+Etoposide,Cytarabine+Daunorubicin, Cytarabine+Doxorubicin, Cytarabine+Idarubicin,Cytarabine+Interferon gamma, Cytarabine+L-asparaginase,Cytarabine+Mitoxantrone, Cytarabine+Fludarabine and Mitoxantrone,Etoposide+Cytarabine, Etoposide+Ifosfamide, Etoposide+Mitoxantrone,Ifosfamide+Etoposide+Mitoxantrone, Ifosfamide+Teniposide,Methotrexate+Mercaptopurine,Methotrexate+Mercaptopurine+Vincristine+Prednisone,Phenylbutyrate+Cytarabine, Phenylbutyrate+Etoposide,Phenylbutyrate+Topotecan, Phenylbutyrate+Tretinoin, Quinine+Doxorubicin,Quinine+Mitoxantrone+Cytarabine, Thioguanine+Cytarabine+Amsacrine,Thioguanine+Etoposide+Idarubicin, Thioguanine+Retinoicacid+Cholecaliferol, Vincristine+Prednisone, Vincristine+Prednisone andL-asparaginase,Vincristine+Dexamethasone/Prednisone+Asparaginase+Daunorubicin/Doxorubicin,Vincristine+Dexamethasone/Prednisone+Asparaginase+Daunorubicin/Doxorubicin+Filgrastim,Vincristine+Dexamethasone/Prednisone+Asparaginase+Daunorubicin/Doxorubicin+Cyclophosphamide+Methotrexate,andVincristine+Dexamethasone/Prednisone+Asparaginase+Daunorubicin/Doxorubicin+Cyclophosphamide+Methotrexate+Filgrastim.

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agent combinations in the treatment, amelioration and/orprevention of acute lymphocytic leukemia.

Compositions of the present invention may be administered in combinationwith one or more therapeutic agents useful in the treatment of chroniclymphocytic leukemia including, but not limited to, Chlorambucil(Leukeran®), Cladribine (2-CdA, Leustatin®), Cyclophosphamide (Cytoxan®,Neosar®, CTX), Cytarabine (Cytosar-U®, ara-C, cytosine arabinoside,DepoCyt®, cytarabine ocfosfate, ara-CMP), Doxorubicin (Adriamycin®,Doxil®, Rubex®), Fludarabine (Fludara®, FAMP), Pentostatin (Nipent®,2-deoxycoformycin), Prednisone and Vincristine (Oncovorin®, Onco TCS®,VCR, Leurocristine®).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agents in the treatment, amelioration and/or prevention ofchronic lymphocytic leukemia.

Further examples of therapeutic agents useful in the treatment ofchronic lymphocytic leukemia which may be administered in combinationwith compositions of the present invention include, but are not limitedto, Alemtuzumab (Campath®), Aminocamptothecin (9-AC,9-Aminocamptothecin, NSC 603071), Aminopterin, Annamycin (AR-522,annamycin LF, Aronex®), Arabinosylguanine (Ara-G, GW506U78,Nelzarabine®, Compound 506U78), Arsenic trioxide (Trisenox®, ATO,Atrivex®), Bryostatin-1 (Bryostatin®, BMY-45618, NSC-339555), CS-682,Dolastatin-10 (DOLA-10, NSC-376128), Filgrastim (Neupogen®, G-CSF,Leukine), Flavopiridol (NSC-649890, HMR-1275), G3139 (Genasense®,GentaAnticode®, Bcl-2 antisense), Irofulven (MGI-114, Ivofulvan,Acylfulvene analogue), MS-209, Phenylbutyrate, Rituximab® (Rituxan®,anti-CD20 MAb), Thalidomide, Theophylline, TNP-470 (AGM-1470,Fumagillin), UCN-01 (7-hydroxystaurosporine) and WHI-P131.

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agents in the treatment, amelioration and/or prevention ofchronic lymphocytic leukemia.

Preferred combinations of therapeutic agents useful in the treatment ofchronic lymphocytic leukemia which may be administered in combinationwith compositions of the present invention include, but are not limitedto, Fludarabine+Prednisone, andCyclophosphamide+Doxorubicin+Vincristine+Prednisone (CHOP).

In preferred embodiments, compositions of the present invention areadministered in combination with one or more of the above-describedtherapeutic agent combinations in the treatment, amelioration and/orprevention of chronic lymphocytic leukemia.

In an additional embodiment, the antibody and antibody compositions ofthe invention are administered in combination with cytokines. Cytokinesthat may be administered with the antibody and antibody compositions ofthe invention include, but are not limited to, GM-CSF, G-CSF, IL2, IL3,IL4, IL5, IL6, IL7, IL10, IL12, IL13, IL15, anti-CD40, CD40L, IFN-alpha,IFN-beta, IFN-gamma, TNF-alpha, and TNF-beta. In preferred embodiments,antibody and antibody compositions of the invention are administeredwith B Lymphocyte Stimulator (e.g., amino acids 134-285 of SEQ IF DNO:3228). In another embodiment, antibody and antibody compositions ofthe invention may be administered with any interleukin, including, butnot limited to, IL-1alpha, IL-1beta, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7,IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17,IL-18, IL-19, IL-20, IL-21, and IL-22. In preferred embodiments, theantibody and antibody compositions of the invention are administered incombination with IL4 and IL10.

In one embodiment, the antibody and antibody compositions of theinvention are administered in combination with one or more chemokines.In specific embodiments, the antibody and antibody compositions of theinvention are administered in combination with an α(CxC) chemokineselected from the group consisting of gamma-interferon inducibleprotein-10 (γIP-10), interleukin-8 (IL-8), platelet factor-4 (PF4),neutrophil activating protein (NAP-2), GRO-α, GRO-β, GRO-γ,neutrophil-activating peptide (ENA-78), granulocyte chemoattractantprotein-2 (GCP-2), and stromal cell-derived factor-1 (SDF-1, or pre-Bcell stimulatory factor (PBSF)); and/or a β(CC) chemokine selected fromthe group consisting of: RANTES (regulated on activation, normal Texpressed and secreted), macrophage inflammatory protein-1 alpha(MIP-1α), macrophage inflammatory protein-1 beta (MIP-1β), monocytechemotactic protein-1 (MCP-1), monocyte chemotactic protein-2 (MCP-2),monocyte chemotactic protein-3 (MCP-3), monocyte chemotactic protein-4(MCP-4) macrophage inflammatory protein-1 gamma (MIP-1γ), macrophageinflammatory protein-3 alpha (MIP-3α), macrophage inflammatory protein-3beta (MIP-3β), macrophage inflammatory protein-4 (MIP-4/DC-CK-1/PARC),eotaxin, Exodus, and I-309; and/or the γ(C) chemokine, lymphotactin.

In another embodiment, the antibody and antibody compositions of theinvention are administered with chemokine beta-8, chemokine beta-1,and/or macrophage inflammatory protein-4. In a preferred embodiment, theantibody and antibody compositions of the invention are administeredwith chemokine beta-8.

In an additional embodiment, the antibody and antibody compositions ofthe invention are administered in combination with an IL-4 antagonist.IL-4 antagonists that may be administered with the antibody and antibodycompositions of the invention include, but are not limited to: solubleIL-4 receptor polypeptides, multimeric forms of soluble IL-4 receptorpolypeptides; anti-IL-4 receptor antibodies that bind the IL-4 receptorwithout transducing the biological signal elicited by IL-4, anti-IL4antibodies that block binding of IL-4 to one or more IL-4 receptors, andmuteins of IL-4 that bind IL-4 receptors but do not transduce thebiological signal elicited by IL-4. Preferably, the antibodies employedaccording to this method are monoclonal antibodies (including antibodyfragments, such as, for example, those described herein).

The invention also encompasses combining the polynucleotides and/orpolypeptides of the invention (and/or agonists or antagonists thereof)with other proposed or conventional hematopoietic therapies. Thus, forexample, the polynucleotides and/or polypeptides of the invention(and/or agonists or antagonists thereof) can be combined with compoundsthat singly exhibit erythropoietic stimulatory effects, such aserythropoietin, testosterone, progenitor cell stimulators, insulin-likegrowth factor, prostaglandins, serotonin, cyclic AMP, prolactin, andtriiodothyzonine. Also encompassed are combinations of the antibody andantibody compositions of the invention with compounds generally used totreat aplastic anemia, such as, for example, methenolene, stanozolol,and nandrolone; to treat iron-deficiency anemia, such as, for example,iron preparations; to treat malignant anemia, such as, for example,vitamin B₁₂ and/or folic acid; and to treat hemolytic anemia, such as,for example, adrenocortical steroids, e.g., corticoids. See e.g.,Resegotti et al., Panminerva Medica, 23:243-248 (1981); Kurtz, FEBSLetters, 14a:105-108 (1982); McGonigle et al., Kidney Int., 25:437-444(1984); and Pavlovic-Kantera, Expt. Hematol., 8(supp. 8) 283-291 (1980),the contents of each of which are hereby incorporated by reference intheir entireties.

Compounds that enhance the effects of or synergize with erythropoietinare also useful as adjuvants herein, and include but are not limited to,adrenergic agonists, thyroid hormones, androgens, hepatic erythropoieticfactors, erythrotropins, and erythrogenins, See for e.g., Dunn, “CurrentConcepts in Erythropoiesis”, John Wiley and Sons (Chichester, England,1983); Kalmani, Kidney Int., 22:383-391 (1982); Shahidi, New Eng. J.Med., 289:72-80 (1973); Urabe et al., J. Exp. Med., 149:1314-1325(1979); Billat et al., Expt. Hematol., 10:133-140 (1982); Naughton etal., Acta Haemat, 69:171-179 (1983); Cognote et al. in abstract 364,Proceedings 7th Intl. Cong. of Endocrinology (Quebec City, Quebec, Jul.1-7, 1984); and Rothman et al., 1982, J. Surg. Oncol., 20:105-108(1982). Methods for stimulating hematopoiesis comprise administering ahematopoietically effective amount (i.e., an amount which effects theformation of blood cells) of a pharmaceutical composition containingpolynucleotides and/or poylpeptides of the invention (and/or agonists orantagonists thereof) to a patient. The polynucleotides and/orpolypeptides of the invention and/or agonists or antagonists thereof isadministered to the patient by any suitable technique, including but notlimited to, parenteral, sublingual, topical, intrapulmonary andintranasal, and those techniques further discussed herein. Thepharmaceutical composition optionally contains one or more members ofthe group consisting of erythropoietin, testosterone, progenitor cellstimulators, insulin-like growth factor, prostaglandins, serotonin,cyclic AMP, prolactin, triiodothyzonine, methenolene, stanozolol, andnandrolone, iron preparations, vitamin B₁₂, folic acid and/oradrenocortical steroids.

In an additional embodiment, the antibody and antibody compositions ofthe invention are administered in combination with hematopoietic growthfactors. Hematopoietic growth factors that may be administered with theantibody and antibody compositions of the invention include, but are notlimited to, LEUKINE™ (SARGRAMOSTIM™) and NEUPOGEN™ (FILGRASTIM™).

In an additional embodiment, the antibody and antibody compositions ofthe invention are administered in combination with fibroblast growthfactors. Fibroblast growth factors that may be administered with theantibody and antibody compositions of the invention include, but are notlimited to, FGF-1, FGF-2, FGF-3, FGF-4, FGF-5, FGF-6, FGF-7, FGF-8,FGF-9, FGF-10, FGF-11, FGF-12, FGF-13, FGF-14, and FGF-15.

Additionally, the antibody and antibody compositions of the inventionmay be administered alone or in combination with other therapeuticregimens, including but not limited to, radiation therapy. Suchcombinatorial therapy may be administered sequentially and/orconcomitantly.

Kits

The invention also provides a pharmaceutical pack or kit comprising oneor more containers filled with one or more of the ingredients of thepharmaceutical compositions of the invention. Optionally associated withsuch container(s) can be a notice in the form prescribed by agovernmental agency regulating the manufacture, use or sale ofpharmaceuticals or biological products, which notice reflects approvalby the agency of manufacture, use or sale for human administration.

The present invention provides kits that can be used in the abovemethods. In one embodiment, a kit comprises an antibody of theinvention, preferably a purified antibody, in one or more containers. Inan alternative embodiment, a kit comprises an antibody fragment thatimmunospecifically binds to B Lymphocyte Stimulator. In a specificembodiment, the kits of the present invention contain a substantiallyisolated B Lymphocyte Stimulator polypeptide as a control. Preferably,the kits of the present invention further comprise a control antibodywhich does not react with B Lymphocyte Stimulator. In another specificembodiment, the kits of the present invention contain a means fordetecting the binding of an antibody to B Lymphocyte Stimulator (e.g.,the antibody may be conjugated to a detectable substrate such as afluorescent compound, an enzymatic substrate, a radioactive compound ora luminescent compound, or a second antibody which recognizes the firstantibody may be conjugated to a detectable substrate). In specificembodiments, the kit may include a recombinantly produced or chemicallysynthesized B Lymphocyte Stimulator. The B Lymphocyte Stimulatorprovided in the kit may also be attached to a solid support. In a morespecific embodiment the detecting means of the above-described kitincludes a solid support to which B Lymphocyte Stimulator is attached.Such a kit may also include a non-attached reporter-labeled anti-humanantibody. In this embodiment, binding of the antibody to B LymphocyteStimulator can be detected by binding of the said reporter-labeledantibody.

In an additional embodiment, the invention includes a diagnostic kit foruse in screening serum containing antigens of the polypeptide of theinvention. The diagnostic kit includes a substantially isolated antibodyspecifically immunoreactive with B Lymphocyte Stimulator, and means fordetecting the binding of B Lymphocyte Stimulator to the antibody. In oneembodiment, the antibody is attached to a solid support. In a specificembodiment, the antibody may be a monoclonal antibody. The detectingmeans of the kit may include a second, labeled monoclonal antibody.Alternatively, or in addition, the detecting means may include alabeled, competing antigen.

In one diagnostic configuration, test serum is reacted with a solidphase reagent having a surface-bound B Lymphocyte Stimulator obtained bythe methods of the present invention. After B Lymphocyte Stimulatorbinds to a specific antibody, the unbound serum components are removedby washing, reporter-labeled anti-human antibody is added, unboundanti-human antibody is removed by washing, and a reagent is reacted withreporter-labeled anti-human antibody to bind reporter to the reagent inproportion to the amount of bound anti-B Lymphocyte Stimulator antibodyon the solid support. Typically, the reporter is an enzyme which isdetected by incubating the solid phase in the presence of a suitablefluorometric, luminescent or colorimetric substrate.

The solid surface reagent in the above assay is prepared by knowntechniques for attaching protein material to solid support material,such as polymeric beads, dip sticks, 96-well plate or filter material.These attachment methods generally include non-specific adsorption ofthe protein to the support or covalent attachment of the protein,typically through a free amine group, to a chemically reactive group onthe solid support, such as an activated carboxyl, hydroxyl, or aldehydegroup. Alternatively, streptavidin coated plates can be used inconjunction with biotinylated antigen(s).

Thus, the invention provides an assay system or kit for carrying outthis diagnostic method. The kit generally includes a support withsurface-bound recombinant B Lymphocyte Stimulator, and areporter-labeled anti-human antibody for detecting surface-bound anti-BLymphocyte Stimulator antibody.

In specific embodiments, the present invention encompasses a singlechain Fv (scFv) having an amino acid sequence of one of SEQ ID NOS: 1 to2128.

In specific embodiments, the present invention encompasses a singlechain Fv (scFv) having an amino acid sequence of one of SEQ ID NOS: 1 to46, 321 to 329, 1563 to 1595, and 1881 to 1908.

In specific embodiments, the present invention encompasses a singlechain Fv (scFv) having an amino acid sequence of one of SEQ ID NOS: 1563to 1880.

In specific embodiments, the present invention encompasses a singlechain Fv (scFv) having an amino acid sequence of one of SEQ ID NOS: 1881to 2128.

In specific embodiments, the present invention encompasses a singlechain Fv (scFv) having an amino acid sequence of one of SEQ ID NOS: 1 to1562.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VH domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128, wherein said antibody orfragment thereof immunospecifically binds B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VH domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 46, 321 to 329, 1563 to 1595,and 1881 to 1908.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VH domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1881 to 2128, and in which saidantibody or fragment thereof immunospecifically binds to themembrane-bound form of B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VH domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1563 to 1880, and in which saidantibody or fragment thereof immunospecifically binds to the solubleform of B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128, wherein said antibody orfragment thereof immunospecifically binds B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 46, 321 to 329, 1563 to 1595,and 1881 to 1908.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1881 to 2128, and in which saidantibody or fragment thereof immunospecifically binds to themembrane-bound form of B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1563 to 1880, and in which saidantibody or fragment thereof immunospecifically binds to the solubleform of B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128, wherein said antibody orfragment thereof immunospecifically binds B Lymphocyte Stimulator andwhich also comprises a VH domain from an scFv having an amino acidsequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128, wherein said antibody orfragment thereof immunospecifically binds B Lymphocyte Stimulator andwhich also comprises a VH domain from an scFv having an amino acidsequence of one of SEQ ID NOS: 1 to 2128.and in which said VL and saidVH domains are derived from the same scFv having an amino acid sequenceof one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the present invention encompasses an antibodyor fragment thereof comprising an amino acid sequence of one of SEQ IDNOS: 2129 to 3227 wherein said antibody or fragment thereofimmunospecifically binds B Lymphocyte Stimulator.

In specific embodiments, the antibody or fragment thereof of theinvention is a whole immunoglobulin molecule.

In specific embodiments, the antibody or fragment thereof of theinvention is a Fab fragment.

In specific embodiments, the antibody or fragment thereof of theinvention is a Fv fragment.

In specific embodiments, the present invention encompasses a chimericprotein comprising the antibody or fragment thereof of the inventioncovalently linked to a heterologous polypeptide.

In specific embodiments, the present invention encompasses a compositioncomprising two or more types of antibodies or fragments or variantsthereof, each of which type immunospecifically binds to B LymphocyteStimulator, and each of which type of antibody or fragment thereofcomprises a VH domain from a different scFv having an amino acidsequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the present invention encompasses a compositioncomprising two or more types of antibodies or fragments or variantsthereof, each of which type immunospecifically binds to B LymphocyteStimulator, and each of which type of antibody or fragment thereofcomprises a VL domain from a different scFv having an amino acidsequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the present invention encompasses a compositioncomprising two or more types of antibodies or fragments or variantsthereof, each of which type immunospecifically binds to B LymphocyteStimulator, and each of which type of antibody or fragment thereofcomprises a VL domain from a different scFv having an amino acidsequence of one of SEQ ID NOS: 1 to 2128 and wherein each type ofantibody or fragment thereof further comprises a VH domain from adifferent scFv having an amino acid sequence of one of SEQ ID NOS: 1 to2128.

In specific embodiments, the present invention encompasses a compositioncomprising two or more types of antibodies or fragments or variantsthereof, each of which type immunospecifically binds to B LymphocyteStimulator, and each of which type of antibody or fragment thereofcomprises a VH CDR3 having an amino acid sequence of one of SEQ ID NOS:3129 to 3227.

In specific embodiments, the present invention encompasses a panel oftwo or more types of antibodies or fragments or variants thereof, eachof which type immunospecifically binds to B Lymphocyte Stimulator, andeach of which type of antibody or fragment thereof comprises a VH domainfrom a different scFv having an amino acid sequence of one of SEQ ID NO:1 to 2128.

In specific embodiments, the present invention encompasses a panel oftwo or more types of antibodies or fragments or variants thereof, eachof which type immunospecifically binds to B Lymphocyte Stimulator, andeach of which type of antibody or fragment thereof comprises a VL domainfrom a different scFv having an amino acid sequence of one of SEQ ID NO:1 to 2128.

In specific embodiments, the present invention encompasses a panel oftwo or more types of antibodies or fragments or variants thereof, eachof which type immunospecifically binds to B Lymphocyte Stimulator, andeach of which type of antibody or fragment thereof comprises a VL domainfrom a different scFv having an amino acid sequence of one of SEQ ID NO:1 to 2128 and wherein each type of antibody or fragment furthercomprises a VH domain from a different scFv having an amino acidsequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the present invention encompasses a panel oftwo or more antibodies or fragments or variants thereof, each of whichtype immunospecifically binds to B Lymphocyte Stimulator, and each ofwhich type of antibody or fragment thereof comprises a VHCDR3 from adifferent scFv having an amino acid sequence of one of SEQ ID NOS: 2129to 3227.

In specific embodiments, the antibodies or fragments thereof of theantibody panel of the invention, are each in a well of a 96 well plate.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VH domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1 to 2128, wherein saidantibody or fragment thereof immunospecifically binds B LymphocyteStimulator.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VH domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1 to 46, 321 to 329, 1563to 1595, and 1881 to 1908, wherein said antibody or fragment thereofimmunospecifically binds B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VH domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1881 to 1908, wherein theantibody of fragment thereof immunospecifically binds the membrane-boundform of B Lymphocyte Stimulator.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VH domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1563 to 1569, wherein saidantibody of fragment thereof immunospecifically binds the soluble formof B Lymphocyte Stimulator. The present invention also encompassesvectors comprising the isolated nucleic acid molecule described above,including vectors comprising a nucleotide sequence which regulates theexpression of the antibody or fragment thereof encoded by theabove-described nucleic acid molecule. Additionally the presentinvention also encompasses host cells, including mammalian host cells,comprising the above-described nucleic acid molecule which is operablylinked to a heterologous promoter, as well as host cells, includingmammalian host cells, comprising the above-described vectors.Additionally, the present invention also provides a method for producingan antibody or fragment thereof comprising culturing the above-describedhost cells under conditions in which the nucleic acid molecule isexpressed.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VL domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1 to 2128, wherein saidantibody or fragment thereof immunospecifically binds B LymphocyteStimulator. The present invention also encompasses vectors comprisingthe isolated nucleic acid molecule described above, including vectorscomprising a nucleotide sequence which regulates the expression of theantibody or fragment thereof encoded by the above-described nucleic acidmolecule. Additionally the present invention also encompasses hostcells, including mammalian host cells, comprising the above-describednucleic acid molecule which is operably linked to a heterologouspromoter, as well as host cells, including mammalian host cells,comprising the above-described vectors. Additionally, the presentinvention also provides a method for producing an antibody or fragmentthereof comprising culturing the above-described host cells underconditions in which the nucleic acid molecule is expressed.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VL domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1 to 46, 321 to 329, 1563to 1595, and 1881 to 1908, wherein said antibody or fragment thereofimmunospecifically binds B Lymphocyte Stimulator. The present inventionalso encompasses vectors comprising the isolated nucleic acid moleculedescribed above, including vectors comprising a nucleotide sequencewhich regulates the expression of the antibody or fragment thereofencoded by the above-described nucleic acid molecule. Additionally thepresent invention also encompasses host cells, including mammalian hostcells, comprising the above-described nucleic acid molecule which isoperably linked to a heterologous promoter, as well as host cells,including mammalian host cells, comprising the above-described vectors.Additionally, the present invention also provides a method for producingan antibody or fragment thereof comprising culturing the above-describedhost cells under conditions in which the nucleic acid molecule isexpressed.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VL domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1881 to 2128, wherein theantibody of fragment thereof immunospecifically binds the membrane-boundform of B Lymphocyte Stimulator. The present invention also encompassesvectors comprising the isolated nucleic acid molecule described above,including vectors comprising a nucleotide sequence which regulates theexpression of the antibody or fragment thereof encoded by theabove-described nucleic acid molecule. Additionally the presentinvention also encompasses host cells, including mammalian host cells,comprising the above-described nucleic acid molecule which is operablylinked to a heterologous promoter, as well as host cells, includingmammalian host cells, comprising the above-described vectors.Additionally, the present invention also provides a method for producingan antibody or fragment thereof comprising culturing the above-describedhost cells under conditions in which the nucleic acid molecule isexpressed.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VL domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1563 to 1880, wherein saidantibody of fragment thereof immunospecifically binds the soluble formof B Lymphocyte Stimulator. The present invention also encompassesvectors comprising the isolated nucleic acid molecule described above,including vectors comprising a nucleotide sequence which regulates theexpression of the antibody or fragment thereof encoded by theabove-described nucleic acid molecule. Additionally the presentinvention also encompasses host cells, including mammalian host cells,comprising the above-described nucleic acid molecule which is operablylinked to a heterologous promoter, as well as host cells, includingmammalian host cells, comprising the above-described vectors.Additionally, the present invention also provides a method for producingan antibody or fragment thereof comprising culturing the above-describedhost cells under conditions in which the nucleic acid molecule isexpressed.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VL domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1 to 2128, wherein saidantibody or fragment thereof immunospecifically binds B LymphocyteStimulator and which also comprises a VH domain from an scFv having anamino acid sequence of one of SEQ ID NOS: 1 to 2128. The presentinvention also encompasses vectors comprising the isolated nucleic acidmolecule described above, including vectors comprising a nucleotidesequence which regulates the expression of the antibody or fragmentthereof encoded by the above-described nucleic acid molecule.Additionally the present invention also encompasses host cells,including mammalian host cells, comprising the above-described nucleicacid molecule which is operably linked to a heterologous promoter, aswell as host cells, including mammalian host cells, comprising theabove-described vectors. Additionally, the present invention alsoprovides a method for producing an antibody or fragment thereofcomprising culturing the above-described host cells under conditions inwhich the nucleic acid molecule is expressed.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VL domain from an scFv havingan amino acid sequence of one of SEQ ID NOS: 1 to 2128, wherein saidantibody or fragment thereof immunospecifically binds B LymphocyteStimulator and which also comprises a VH domain from an scFv having anamino acid sequence of one of SEQ ID NOS: 1 to 2128 and in which said VLdomain and said VH domain are derived from the same scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128. The present inventionalso encompasses vectors comprising the isolated nucleic acid moleculedescribed above, including vectors comprising a nucleotide sequencewhich regulates the expression of the antibody or fragment thereofencoded by the above-described nucleic acid molecule. Additionally thepresent invention also encompasses host cells, including mammalian hostcells, comprising the above-described nucleic acid molecule which isoperably linked to a heterologous promoter, as well as host cells,including mammalian host cells, comprising the above-described vectors.Additionally, the present invention also provides a method for producingan antibody or fragment thereof comprising culturing the above-describedhost cells under conditions in which the nucleic acid molecule isexpressed.

In specific embodiments, the present invention encompasses an isolatednucleic acid molecule comprising a nucleotide sequence encoding anantibody or fragment thereof comprising a VHCDR3 from an scFv having anamino acid sequence of one of SEQ ID NOS: 2129 to 3227, wherein saidantibody or fragment thereof immunospecifically binds B LymphocyteStimulator. The present invention also encompasses vectors comprisingthe isolated nucleic acid molecule described above, including vectorscomprising a nucleotide sequence which regulates the expression of theantibody or fragment thereof encoded by the above-described nucleic acidmolecule. Additionally the present invention also encompasses hostcells, including mammalian host cells, comprising the above-describednucleic acid molecule which is operably linked to a heterologouspromoter, as well as host cells, including mammalian host cells,comprising the above-described vectors. Additionally, the presentinvention also provides a method for producing an antibody or fragmentthereof comprising culturing the above-described host cells underconditions in which the nucleic acid molecule is expressed.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a VH domain encoded by a nucleotide sequence that hybridizesunder stringent conditions to a nucleotide sequence encoding a VH domainfrom an scFv having an amino acid sequence of one of SEQ ID NOS: 1 to2128.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a VL domain encoded by a nucleotide sequence that hybridizesunder stringent conditions to a nucleotide sequence encoding a VL domainfrom an scFv having an amino acid sequence of one of SEQ ID NOS: 1 to2128.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a VH domain encoded by a nucleotide sequence that hybridizesunder highly stringent conditions to a nucleotide sequence encoding a VHdomain from an scFv having an amino acid sequence of one of SEQ ID NOS:1 to 2128.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a VL domain encoded by a nucleotide sequence that hybridizesunder highly stringent conditions to a nucleotide sequence encoding a VLdomain from an scFv having an amino acid sequence of one of SEQ ID NOS:1 to 2128.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a CDR encoded by a nucleotide sequence that hybridizes understringent conditions to a nucleotide sequence encoding a CDR from anscFv having an amino acid sequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a CDR encoded by a nucleotide sequence that hybridizes underhighly stringent conditions to a nucleotide sequence encoding a CDR froman scFv having an amino acid sequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a VH CDR3 encoded by a nucleotide sequence that hybridizesunder stringent conditions to a nucleotide sequence encoding a VH CDR3having an amino acid sequence of one of SEQ ID NOS: 2129 to 3227.

In specific embodiments, the present invention provides an antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator, said antibody or fragment thereof comprising an amino acidsequence of a VH CDR3 encoded by a nucleotide sequence that hybridizesunder highly stringent conditions to a nucleotide sequence encoding a VHCDR3 having an amino acid sequence of one of SEQ ID NOS: 2129 to 3227.

In specific embodiments, the present invention provides a method fordetecting of aberrant expression of B Lymphocyte Stimulator, comprising:

assaying the level of B Lymphocyte Stimulator expression in cells or atissue sample of an individual using one or more antibodies or fragmentsor variants thereof that immunospecifically bind B LymphocyteStimulator; and

comparing the level of B Lymphocyte Stimulator assayed in the cells or atissue sample with a standard level of B Lymphocyte Stimulator or alevel of B Lymphocyte Stimulator in cells or a tissue sample from anindividual without aberrant B Lymphocyte Stimulator expression, whereinan increase or decrease in the assayed level of B Lymphocyte Stimulatoror level in cells or a tissue sample from an individual without aberrantB Lymphocyte Stimulator expression compared to the standard level of BLymphocyte Stimulator is indicative of aberrant expression.

In specific embodiments, the present invention provides a method fordiagnosing a disease or disorder associated with aberrant B LymphocyteStimulator expression or activity, comprising:

administering to a subject an effective amount of a labeled antibody orfragment thereof that immunospecifically binds to B LymphocyteStimulator;

waiting for a time interval following the administering for permittingthe labeled antibody or fragment thereof to preferentially concentrateat sites in the subject where B Lymphocyte Stimulator is expressed;

determining background level; and

detecting the labeled antibody or fragment thereof in the subject, suchthat detection of labeled antibody or fragment thereof above thebackground level indicates that the subject has a particular disease ordisorder associated with aberrant expression of B Lymphocyte Stimulator.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above comprises a VH domain froman scFv having an amino acid sequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above comprises a VL domain froman scFv having an amino acid sequence of one of SEQ ID NOS: 1 to 2128.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above comprises a VH CDR3 havingan amino acid sequence of one of SEQ ID NOS: 2129 to 3227.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above is conjugated to adiagnostic agent.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above is conjugated to adiagnostic agent wherein the diagnostic agent is horseradish peroxidase,alkaline phosphatase, beta-galactosidase, or acetylcholinesterase.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above is conjugated to adiagnostic agent wherein the diagnostic agent is fluorescein,fluorescein isothiocyanate, rhodamine, dichlorotriazinylaminefluorescein, dansyl chloride or phycoerythrin.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above is conjugated to adiagnostic agent wherein the diagnostic agent is ¹²⁵I, ¹³¹I, ¹¹¹In, ⁹⁰Yor ⁹⁹Tc.

In specific embodiments, the antibody or fragment thereof utilized inthe two methods described immediately above is conjugated to adiagnostic agent wherein the diagnostic agent is luciferase, luciferinor aequorin.

A pharmaceutical composition comprising at least one antibody orfragment thereof of comprising a VH domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128, wherein said antibody orfragment thereof immunospecifically binds B Lymphocyte Stimulator and apharmaceutically acceptable carrier.

A pharmaceutical composition comprising at least one antibody orfragment thereof of comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128, wherein said antibody orfragment thereof immunospecifically binds B Lymphocyte Stimulator and apharmaceutically acceptable carrier.

A pharmaceutical composition comprising at least one antibody orfragment thereof of comprising a VL domain from an scFv having an aminoacid sequence of one of SEQ ID NOS: 1 to 2128, wherein said antibody orfragment thereof immunospecifically binds B Lymphocyte Stimulator andwhich also comprises a VH domain from an scFv having an amino acidsequence of one of SEQ ID NOS: 1 to 2128 and a pharmaceuticallyacceptable carrier.

A pharmaceutical composition comprising at least one antibody orfragment thereof of comprising an amino acid sequence of one of SEQ IDNOS: 2129 to 3227 wherein said antibody or fragment thereofimmunospecifically binds B Lymphocyte Stimulator and a pharmaceuticallyacceptable carrier.

A method of treating, preventing or ameliorating a disease or disorderassociated with aberrant B Lymphocyte Stimulator expression or activity,comprising administering to an animal in need thereof the pharmaceuticalcomposition comprising at least one antibody or fragment thereof ofcomprising a VL domain from an scFv having an amino acid sequence of oneof SEQ ID NOS: 1 to 2128, wherein said antibody or fragment thereofimmunospecifically binds B Lymphocyte Stimulator and which alsocomprises a VH domain from an scFv having an amino acid sequence of oneof SEQ ID NOS: 1 to 2128 and a pharmaceutically acceptable carrier in anamount effective to treat, prevent or ameliorate the disease ordisorder. This method may be used to treat an infectious disorder,cancer, and/or an autoimmune disease such as lupus or glomerularnephritis.

A method of treating, preventing or ameliorating a disease or disorderassociated with aberrant B Lymphocyte Stimulator expression or activity,comprising administering to an animal in need thereof the pharmaceuticalcomposition comprising at least one antibody or fragment thereof ofcomprising a VL domain from an scFv having an amino acid sequence of oneof SEQ ID NOS: 1 to 2128, wherein said antibody or fragment thereofimmunospecifically binds B Lymphocyte Stimulator and a pharmaceuticallyacceptable carrier in an amount effective to treat, prevent orameliorate the disease or disorder. This method may be used to treat aninfectious disorder, cancer, and/or an autoimmune disease such as lupusor glomerular nephritis.

A method of treating, preventing or ameliorating a disease or disorderassociated with aberrant B Lymphocyte Stimulator expression or activity,comprising administering to an animal in need thereof the pharmaceuticalcomposition comprising at least one antibody or fragment thereof ofcomprising a VL domain from an scFv having an amino acid sequence of oneof SEQ ID NOS: 1 to 2128, wherein said antibody or fragment thereofimmunospecifically binds B Lymphocyte Stimulator and which alsocomprises a VH domain from an scFv having an amino acid sequence of oneof SEQ ID NOS: 1 to 2128 and a pharmaceutically acceptable carrier in anamount effective to treat, prevent or ameliorate the disease ordisorder. This method may be used to treat an infectious disorder,cancer, and/or an autoimmune disease such as lupus or glomerularnephritis.

A method of treating, preventing or ameliorating a disease or disorderassociated with aberrant B Lymphocyte Stimulator expression or activity,comprising administering to an animal in need thereof the pharmaceuticalcomposition of comprising at least one antibody or fragment thereof ofcomprising an amino acid sequence of one of SEQ ID NOS: 2129 to 3227wherein said antibody or fragment thereof immunospecifically binds BLymphocyte Stimulator and a pharmaceutically acceptable carrier in anamount effective to treat, prevent or ameliorate the disease ordisorder. This method may be used to treat an infectious disorder,cancer, and/or an autoimmune disease such as lupus or glomerularnephritis.

This method may be used to treat an infectious disorder, cancer, and/oran autoimmune disease such as lupus or glomerular nephritis.

EXAMPLES Abbreviations

-   0.2 M Tris-HCl, 0.5 mM EDTA, 0.5 M sucrose (TES)-   1-ethyl-3-[3-dimethylaminopropyl]carbo diimide hydrochloride (EDC)-   2TY supplemented with 100 μg/ml ampicillin and 2% glucose (2TYAG)-   2TY supplemented with 100 μg/ml ampicillin and 50 μg/mlkanamycin    (2TYAK)-   3,3′,5,5′-Tetramethyl Benzidine (TMB)-   50% inhibitory concentration (IC₅₀)-   6×PBS containing 18% Marvel blocking solution (6×MPBS)-   Absorbance (A)-   Bovine serum albumin (BSA)-   Enzyme linked immunosorbent assay (ELISA)-   Foetal calf serum (FCS)-   Heavy chain variable (V_(H))-   Hepes buffered saline (HBS)-   Horseradish peroxidase (HRP)-   Immobilised Metal Affinity Chromatography (IMAC)-   Isopropyl β-D-thiogalactopyranoside (IPTG)-   Light chain variable(V_(L))-   Multiplicity of infection (MOI)-   N-[2-hydroxyethyl]piperazine-N′-[2-ethanesulfonic acid] (Hepes)-   Nanomolar (nM)-   N-Hydroxysuccinimide (NHS)-   PBS containing 3% Marvel (MPBS)-   Phosphate Buffered Saline (PBS)-   Phosphate Buffered Saline+0.1% (v/v) Tween 20 (PBST)-   Picomolar (pM)-   Single chain fragment variable (scFv)-   Tumour Necrosis Factor-alpha (TNF-α)-   Tumour Necrosis Factor-beta (TNF-β)-   TNF-related apoptosis inducing ligand (TRAIL)

Definitions:

In the following section “immobilized B Lymphocyte Stimulator” refers toa soluble form of B Lymphocyte Stimulator or biotinylated B LymphocyteStimulator coated on a plastic assay plate (e.g., a 96 well plate), butdoes not refer to histidine tagged B Lymphocyte Stimulator coated on aplastic assay plate.; “biotinylated B Lymphocyte Stimulator” is asoluble form of B Lymphocyte Stimulator except when used to coat anELISA plate, in which case it would be “immobilized B LymphocyteStimulator.” Membrane bound forms of B Lymphocyte Stimulator include,but are not limited to, U937 and P388 plasma membranes. Antibodies ofthe present invention are defined as able to bind the membrane boundand/or soluble forms of B Lymphocyte Stimulator according to the assaysdescribed in Examples 1 through 19.

Example 1 Antibodies Immunospecifically Binding to Soluble AndMembrane-Bound B Lymphocyte Stimulator

A library of phage was screened in an assay to identify those phagedisplaying scFvs that immunospecifically bind to the soluble andmembrane-bound forms of B Lymphocyte Stimulator. Phage displaying scFvsthat bound to immobilized B Lymphocyte Stimulator were identified afterpanning on immobilized B Lymphocyte Stimulator and assessment by ELISAfor binding to immobilized B Lymphocyte Stimulator. The B LymphocyteStimulator that was immobilized on plates for these assays was purifiedfrom supernatants of Sf9 cells infected with a baculovirus expressionconstruct as described in Moore et al., Science 285:260-263 which ishereby incorporated by reference in its entirety. Each of the identifiedscFvs were then sequenced. Certain sequences were isolated multipletimes, thus a panel (panel 1) containing one member of each uniquesequences was generated and further characterized for their ability toimmunospecifically bind to the soluble and membrane-bound forms of BLymphocyte Stimulator.

The derived amino acid sequences of these scFvs are shown in Table 1above. The individual V_(H) and V_(L) segments of the scFvs were alignedto the known human germline sequences in V-BASE (Tomlinson et al, whichcan accessed on the United Kingdom Medical Research Council (MRC) Centrefor Protein Engineering website) and the closest germline identified.

Example 2 Specificity of scFvs for B Lymphocyte Stimulator andMembrane-Bound B Lymphocyte Stimulator

The specificity of each of the scFvs for both B Lymphocyte Stimulatorand membrane-bound B Lymphocyte Stimulator was determined by phageELISA. B Lymphocyte Stimulator was immobilised onto plastic as apurified soluble form of the protein or as a membrane-bound form presenton plasma membrane preparations from the human macrophage-like cellline, U937.

Maintenance of U937 Cells

U937 cells are a human monocyte-like, histiocytic lymphoma cell lineknown to express B Lymphocyte Stimulator on their plasma membranes. Theywere maintained in RPMI-1640 supplemented with 4 mM L-glutamine, 10%FCS, 10 U penicillin, 100 g/ml streptomycin (all reagents from Sigma).The cells were thawed from frozen stock and are either used for plasmamembrane preparation, or split 1:5, after 2 days in culture when thecell density reaches 1×10⁶/ml.

Preparation of U937 Plasma Membranes

To prepare plasma membranes, 1×10⁹ U937 cells were harvested from theirculture medium by centrifugation at 1000 rpm at 4° C. for 5 minutes in abenchtop centrifuge. The cells were resuspended in 40 ml 12 mM Tris, pH7.5, 250 mM sucrose and placed on ice. The cells are then lysed using ahand-held electric homogenizer (Labortechnik IKA Ultra-Turrax) for four,one minute, bursts. To check that cell lysis had occurred, 10 μl celllysate was added to 10 μl Trypan blue and the cell lysate was examinedunder a microscope. After confirming lysis, the homogenate wascentrifuged at 270×g, for 10 minutes at 4° C. to pellet the nuclearfraction and the supernatant was retained. The supernatant wascentrifuged at 8000×g, 10 mins, 4° C., to pellet the mitochondrial andlysosomal fractions and the supernatant was retained. The supernatantwas then centrifuged at 100000×g, 60 mins, 4° C. to pellet the plasmamembrane enriched fraction. The supernatant was discarded and the plasmamembrane pellet was resuspended in 1 ml PBS and stored at −70° C. Theprotein concentration of the plasma membrane fraction was determinedusing a protein quantification kit (Biorad). Typical yields were between5 and 10 mg of plasma membranes.

Phage ELISA

To determine the specificity of each of the unique scFvs, a phage ELISAwas performed for each scFv against human B Lymphocyte Stimulator, U937plasma membranes, TNFα (R&D Systems, Minneapolis, Minn.), BSA anduncoated well. Individual E. coli colonies containing a phagemidrepresenting one of the unique scFvs from panel lwere inoculated into96-well plates containing 100 μl 2TYAG medium per well. Plates wereincubated at 37° C. for 4 hours, shaking. M13KO7 helper phage was addedto each well to a MOI of 10 and the plates were incubated for a further1 hour at 37° C. The plates were centrifuged in a benchtop centrifuge at2000 rpm for 10 minutes. The supernatant was removed and cell pelletswere resuspended in 100 μl TYAK and incubated at 30° C. overnight,shaking. The next day, plates were centrifuged at 2000 rpm for 10 minand the 100 μl phage-containing supernatant from each well carefullytransferred into a fresh 96-well plate. Twenty μl of 6×MPBS was added toeach well, and incubated at room temperature for 1 hour to pre-block thephage prior to ELISA.

Flexible 96-well plates (Falcon) were coated overnight at 4° C. withhuman B Lymphocyte Stimulator (1 μg/ml) in PBS, U937 plasma membranes(10 μg/ml) in PBS, TNFα (1 μg/ml) in PBS, BSA (1 μg/ml) in PBS, or PBS.After coating, the solutions were removed from the wells, and the plateswere blocked for 1 hour at room temperature in MPBS. The plates werewashed 3 times with PBS and then 50 μl of pre-blocked phage was added toeach well. The plates were incubated at room temperature for 1 hour andthen washed with 3 changes of PBST followed by 3 changes of PBS. To eachwell, 50 μl of an anti-gene VIII-HRP conjugate (Pharmacia) at a 1 to5000 dilution in MPBS was added and the plates incubated at roomtemperature for 1 hour. Each plate was washed three times with PBSTfollowed by three times with PBS. Then 50 μl of an HRP-labelledanti-mouse polymer (DAKO EnVision) diluted 1/50 in 3% MPBS was added andincubated for 1 hour at room temperature. Each plate was then washedthree times with PBST followed by three times with PBS. Fifty μl of TMBsubstrate was then added to each well, and incubated at room temperaturefor 30 minutes or until colour development. The reaction was stopped bythe addition of 25 μl of 0.5 M H₂SO₄. The signal generated was measuredby reading the absorbance at 450 nm (A₄₅₀) using a microtiter platereader (Bio-Rad 3550).

The results for 3 clones (I006E07, I008D05 and I016F04) are shown inFIG. 1. All 3 scFvs recognize immobilized B Lymphocyte Stimulator andU937 plasma membranes but do not recognize TNFα, BSA or an uncoated well(PBS only). These results indicate that these scFvs specificallyrecognize immobilized B Lymphocyte Stimulator and membrane-bound BLymphocyte Stimulator.

Example 3 Inhibition in an In Vitro Receptor Binding Assay by PhageScFvs

All of the unique phage scFvs in panel 1 were assessed for their abilityto inhibit soluble B Lymphocyte Stimulator binding to its cognatereceptor on IM9 cells.

Biotinylation of B Lymphocyte Stimulator

One hundred μg of either human or mouse B Lymphocyte Stimulator wasdialysed overnight at 4° C. against 50 mM sodium bicarbonate (sodiumhydrogen carbonate) pH8.5 using a slide-a-lyzer cassette (Pierce). Thenext day, NHS-biotin (Pierce) was dissolved in DMSO to 13.3 mg/ml. Thiswas then added to the B Lymphocyte Stimulator at a molar ratio of 20:1biotin:B Lymphocyte Stimulator, mixed and incubated on ice for 2 hours.The biotinylated B Lymphocyte Stimulator was then dialysed back intosterile PBS (Sigma) using a slide-a-lyzer cassette overnight at 4° C.The biological activity of the biotinylated B Lymphocyte Stimulator wasconfirmed using the receptor binding inhibition assay (see below).

Maintenance of IM9 Cells

IM9 cells are a human B lymphocyte cell line. They were maintained inRPMI-1640 supplemented with 4 mM L-glutamine, 10% FCS, 10 U penicillin,100 g/ml streptomycin (all reagents from Sigma). The cells are thawedfrom frozen stock and can be used in assays after 5 days in culture whenthey reach a density of 4-8×10⁵ /ml.

Receptor Binding Inhibition Assay

Individual E. coli colonies containing a phagemid representing one ofthe unique scFvs from panel 1 were inoculated into 96-well platescontaining 100 μl 2TYAG medium per well. Plates were incubated at 37° C.for 4 hours, shaking. M13KO7 helper phage was added to each well to aMOI of 10 and the plates were incubated for a further 1 hour at 37° C.The plates were centrifuged in a benchtop centrifuge at 2000 rpm for 10minutes. The supernatant was removed and cell pellets were resuspendedin 100 μl 2TYAK and incubated at 30° C. overnight, shaking. The nextday, plates were centrifuged at 2000 rpm for 10 min and the 100 μlphage-containing supernatant from each well carefully transferred into afresh 96-well plate. Phage were diluted 1 in 2 in MPBS prior to use.

Flat-bottomed 96-well plates (Costar) were coated with 100 μl per wellof a 1:10 dilution of poly-L-lysine (Sigma) in PBS for 1 hour at roomtemperature. The plates were then washed twice with water, allowed toair-dry and placed at 4° C. overnight. One hundred μl of IM9 cells (at10⁶/ml in RPMI-1640 culture medium) were then added to each well. Plateswere then centrifuged at 3200 rpm for 5 mins to pellet the cells. Themedia was carefully aspirated and 200 μl of MPBS added to each well. Theplates were then allowed to block for 1 hour at room temperature.

To a separate 96-well plate 10 μl of biotinylated B LymphocyteStimulator (at 162.5 ng/ml) in MPBS was added to each well to give afinal concentration of 25 ng/ml. Fifty-five μl of each appropriate phagesupernatant was added to each well and the final volume in each well was65 μl. Plates were then incubated at room temperature for 30 minutes.

The IM9 coated plates were washed twice in PBS, tapped dry andimmediately 50 μl of the phage/biotinylated-B Lymphocyte Stimulator mixwas added and incubated at room temperature for 1 hour. Plates werewashed three times in PBST and three times in PBS, tapped dry and 50 μlof streptavidin-Delfia (Wallac) was added to each well at 1:1000dilution in the Manufacturer's assay buffer. The plates were thenincubated at room temperature for 1 hour and washed six times in Delfiawash solution (Wallac). After tapping the plates dry, 100 μl per well ofDelfia enhancement solution (Wallac) was added. The plates were gentlytapped to encourage micelle formation, incubated at room temperature for10 minutes, and fluorescence read on a Wallac 1420 workstation at 6520nM.

Results for 3 phage scFvs (I001C09, I018D07 and I016H07) that inhibitedthe binding of biotinylated B Lymphocyte Stimulator are shown in FIG. 2.Maximal binding of biotinylated B Lymphocyte Stimulator to its receptor(bio-B Lymphocyte Stimulator only), the background signal in the absenceof biotinylated B Lymphocyte Stimulator (no bio-B LymphocyteStimulator), and results with an irrelevant (i.e., does not recognize BLymphocyte Stimulator) phage antibody are also shown. All 3 phage scFvsinhibited biotinylated B Lymphocyte Stimulator binding to its receptoron IM9 cells, identifying these scFvs as scFvs that bind the solubleform of B Lymphocyte Stimulator. These scFvs also bind to U937membranes, thus they also bind the membrane bound form of B LymphocyteStimulator.

Forty-eight of the scFvs from panel 1 that demonstrated the greatestinhibition as phage particles in this assay were chosen for furtherstudy. These 48 scFvs are listed in Table 3.

TABLE 3 scFvs that Inhibit the Binding of Biotinylated-B LymphocyteStimulator to its Receptor Antibody Antibody Antibody Antibody AntibodyI008C02 I029D07 I008C03 I008C12 I028A06 I022E02 I061E07 I007H08 I061H011031C03 I018C02 I006D07 I008A11 I006D08 I031F02 I008B01 I017D10 I061D02I026E03 I031F09 I016F04 I007B03 I008A09 I027A07 I031G11 I016E05 I018C10I007F11 I016H07 I050A07 1018H08 I001C09 I037E07 I021B05 I050A12 I018H09I018D07 I037E12 I031G10 I050B11 I029F11 I016F02 I031G08 I051C04 I022D01I031C07 I003F12 I012A06

Example 4 Specificity of Anti-B Lymphocyte Stimulator Antibodies

The specificity of the 48 scFvs listed in Table 3 for human and murine BLymphocyte Stimulator was determined using phage ELISA.

Phage ELISA

To determine the specificity of the 48 scFvs, a phage ELISA wasperformed against human and mouse B Lymphocyte Stimulator, and a panelof related and unrelated human antigens: Fas ligand, TRAIL, TNFα, TNFβ,and PBS. The: Fas ligand, TRAIL, TNFα, and TNFβ antigens were obtainedfrom R&D Systems, Minneapolis, Minn. Individual E. coli coloniescontaining phagemid were inoculated into 5 ml 2YTAG and incubated at 37°C. for 4 hours, shaking. M13KO7 helper phage (Pharmacia) was added toeach tube to a MOI of 10 and incubated for 30 minutes at 37° C. for 1hour, the first 30 minutes static and the final 30 minutes with gentleshaking. Cells were pelleted by centrifugation at 3,500 rpm for 10minutes and the supernatant discarded. Cell pellets were resuspended in5 ml 2TYAK and incubated at 30° C. overnight with shaking. The next day,the cells were pelleted by centrifugation at 3,500 rpm for 10 minutes.The phage-containing supernatant (5 ml) was carefully transferred to afresh tube, 1 ml of 6MPBS was added, and the tube was incubated at roomtemperature for 1 hour to pre-block the phage prior to ELISA.

All antigens were coated at 1 μg/ml. ELISAs were performed essentiallyas described in Example 2. The only exception to this being thedetection of phage antibody binding to mouse B Lymphocyte Stimulatorwhere the step involving incubation with the HRP-labelled anti-mousepolymer was omitted. Binding to mouse B Lymphocyte Stimulator wasdetected with TMB as in Example 2.

All 48 scFvs are specific for immobilized human B Lymphocyte Stimulatorand 43 out of the 48 scFvs cross-react with immobilized mouse BLymphocyte Stimulator but not with any other unrelated or relatedantigen tested. I008CO3, I007F11, I037E07, I037E12, and I016H07 did notbind murine B Lymphocyte Stimulator. Results for two scFvs, I022D01 andI031F02, are shown in FIG. 3. Both these scFvs specifically recognizehuman and mouse B Lymphocyte Stimulator but not any other unrelated orrelated antigen tested.

Example 5 Specificity for the Membrane-Bound Form of B LymphocyteStimulator

The specificity of 48 scFvs for membrane-bound B Lymphocyte Stimulatorwas determined by the phage ELISA described in Example 2. B LymphocyteStimulator was immobilised onto plastic as a membrane-bound form presenton plasma membranes preparations from the human macrophage-like cellline, U937. This cell line is known to express the membrane-bound formof human B Lymphocyte Stimulator.

To demonstrate that this binding is specific for membrane-bound BLymphocyte Stimulator, a competition ELISA was developed to determine ifthe ELISA signal for an individual antibody on U937′s could be competedout by pre-incubation with either B Lymphocyte Stimulator or TNFα. Ananti-B Lymphocyte Stimulator antibody that also recognizesmembrane-bound B Lymphocyte Stimulator would be expected to demonstratea signal reduction with free B Lymphocyte Stimulator but not free TNFα.

Competition ELISA

Individual E. coli colonies containing phagemid for each of the 48 scFvslisted in Table 3 were inoculated into 5 ml 2YTAG and incubated at 37°C. for 4 hours, shaking. M13KO7 helper phage (Pharmacia) was added toeach tube to a MOI of 10 and incubated for 30 minutes at 37° C. for 1hour, the first 30 minutes static and the final 30 minutes with gentleshaking. Cells were pelleted by centrifugation at 3,500 rpm for 10minutes and the supernatant discarded. Cell pellets were resuspended in5 ml 2TYAK and incubated at 30° C. overnight with shaking. The next day,the cells were pelleted by centrifugation at 3,500 rpm for 10 minutes.The phage-containing supernatants (5 ml) were carefully transferred to afresh tube.

For each of the 48 scFvs listed in Table 3, two aliquots of 20 μl 6×MPBSwere pipetted into separate wells of a 96-well plate (Greiner). Thefirst aliquot was supplemented with B Lymphocyte Stimulator to a finalconcentration of 0.5 μg/ml. The second aliquot was supplemented withTNF-α to a final concentration of 0.5 μg/ml. Each experiment wasperformed in triplicate. One hundred μl of each phage supernatant wasthen added to each aliquot and mixed by pipetting up and down. The phagewere incubated (±competing antigen) at room temperature for 1 hour.

Flexible 96-well plates (Falcon) were coated overnight at 4° C. with 50μl of 10 μg/ml U937 plasma membranes. After coating, the plates werewashed 3 times with PBS and blocked for 1 hour at room temperature with200 μl MPBS. The plates were washed 3 times with PBS and 50 μl of phage(±competing antigen) was added to each appropriate well. The plates wereincubated at room temperature for 1 hour and then washed with 3 changesof PBST followed by 3 changes of PBS. To each well, 50 μl of a mouseanti-gene VIII-HRP conjugate (Pharmacia) at a 1:5000 dilution in MPBSwas added and the plates incubated at room temperature for 1 hour. Eachplate was washed three times with PBST followed by three times with PBS.Then 50 μl of an HRP-labelled anti-mouse polymer (DAKO EnVision) diluted1:50 in 3% MPBS was added and incubated for 1 hour at room temperature.Each plate was then washed three times with PBST followed by three timeswith PBS. Fifty μl of TMB substrate was then added to each well, andincubated at room temperature for 30 to 60 minutes or until colordevelopment. The reaction was stopped by the addition of 25 μl of 0.5 MH₂SO₄. The signal generated was measured by reading the absorbance at450 nm (A₄₅₀) using a microtiter plate reader (Bio-Rad 3550).

All 48 scFvs bind to U937 plasma membrane preparations. This signalcould be competed out by pre-incubation of the phage antibody with BLymphocyte Stimulator but not by pre-incubation with TNF-α. Thisindicates that the 48 scFvs specifically recognize membrane-bound BLymphocyte Stimulator as well as soluble B Lymphocyte Stimulator.Typical results are exemplified by scFvs I031F09, I050A12 and I051C04and are shown in FIG. 4. All 3 scFvs demonstrate binding to U937 plasmamembranes. This binding was specifically competed out with B LymphocyteStimulator but did not compete with TNF-α, demonstrating specificrecognition of membrane-bound B Lymphocyte Stimulator.

Example 6 scFv Off-Rate Determinations

All off-rate determinations were performed on BIAcore 2000 machines,using the BIAcore 2000 Control Software and evaluated using theBIAevaluation 3.0 software.

Preparation of a Low Density B Lymphocyte Stimulator Surface

A 500RU surface was prepared for kinetic studies with purified scFvs. Alow density B Lymphocyte Stimulator surface (500 RU B LymphocyteStimulator coupled) was prepared in flow cell 2 by amine coupling to aCM5 chip. A new CM5 chip was inserted into the BIAcore and a sensorgraminitiated with HBS buffer at a flow rate of 5 μl/min. The NHS and EDCcoupling solutions (BIAcore) were mixed according to manufacturer'sinstructions and 30 μl injected over the CM5 surface. Fifty μl of BLymphocyte Stimulator at 1 μg/ml in 10 mM sodium acetate buffer, pH4,was then injected followed by 30 μl of ethanolamine-HCl solution(BIAcore). The flow rate was then adjusted to 20 μl/min and 10 μl of 4Mguanidine hydrochloride in HBS injected over the surface. This stripsthe surface of non-covalently bound B Lymphocyte Stimulator.

Measurement of scFv Off-Rate Kinetics on the Low Density Surfaces

The chip containing the low density B Lymphocyte Stimulator surface wasinserted in to the BIAcore. A dilution series of purified scFvs wasprepared in HBS, typically 50 μg/ml doubling dilutions down to 1.5ug/ml. The dilution series was then injected sequentially over the lowdensity B Lymphocyte Stimulator surface (and blank control) using thefollowing program:

MAIN FLOWCELL 1,2,3,4 APROG genab r1d1 ab1 APROG genab r1d2 ab2 APROGgenab r1d3 ab3 APROG genab r1d4 ab4 APROG genab r1d5 ab5 APROG genabr1d6 ab6 APPEND CONTINUE END DEFINE APROG genab PARAM %Abpos %AbId FLOW20 KINJECT %Abpos 200 80 INJECT r1c6 10!guanidine hydrochlorideregeneration step EXTRACLEAN END

Bound scFvs were removed by injecting 10 μl 4MGuHCl in HBS over thesurface between scFv samples.

The binding curves for individual scFvs were analyzed using theBIAevaluation software to determine antibody off-rates. Kinetic analysisfor a typical scFv antibody, I003C02, is shown in FIG. 5. I003C02 has aK_(off)=6×10⁻³ s⁻¹.

Example 7 Inhibition in an In Vitro Receptor Binding Assay by scFvAntibodies

The 48 scFvs listed in Table 3 were purified and assessed for theirability to inhibit B Lymphocyte Stimulator binding to its receptor onIM9 cells.

Purification of scFv

To determine the inhibitory potency of anti-B Lymphocyte StimulatorscFv, scFv's were first prepared by IMAC. 2TYAG (5 ml) was inoculatedwith a single colony and grown overnight at 30° C., shaking. Thisovernight culture was then used to inoculate 500 ml of 2TY containing100 μg/ml ampicillin and 0.1% Glucose, and grown at 30° C., shaking,until an A₆₀₀ of 1.0 was attained. IPTG was added to 1 mM and theculture was grown for a further 3.5 hours at 30° C.

Cells were harvested by centrifugation at 5,000 rpm, and resuspended in10 ml of TES. A further 15 ml of a 1:5 dilution (in water) of TES wasadded, and the cell suspension incubated on a turning wheel at 4° C. for30 minutes. This causes osmotic shock and yields a periplasmic extractcontaining the scFv. Residual cells and debris were pelleted bycentrifugation at 9,000 rpm for 20 minutes at 4° C. The supernatant wastransferred to a new tube, and 50 μl of 1 M MgCl₂ added. Two ml of aNi-NTA agarose (Qiagen), pre-washed with buffer (50 mM sodium phosphate,pH 8, 300 mM NaCl) together with a protease inhibitor tablet (BoehringerMannheim) were then added to the periplasmic extract. The preparationwas incubated, rotating, overnight at 4° C. The Ni-NTA was pelleted bycentrifugation at 2,000 rpm for 5 minutes, and the supernatant wasaspirated. The agarose beads were washed 3 times with 50 ml wash buffer,centrifuging to collect the agarose in between each wash. Ten ml of washbuffer was added after the final wash, and the slurry was loaded on to apolyprep column (BioRad). Two ml elution buffer (50 mM NaPi (sodiumphosphate), pH 8, 300 mM NaCl, 250 mM imidazole) was added to thedrained agarose, and the eluate was collected. IMAC purified scFv wasbuffer exchanged in to PBS by use of a Nap 5 column (Pharmacia)according to the manufacturer's instructions. The A₂₈₀ was read and theprotein concentration determined using a molar extinction coefficient of1 mg/ml protein=A₂₈₀ 1.4. Purified scFv was stored in 500 μl aliquots at−70° C.

Receptor Binding Inhibition Assay

Flat-bottomed 96-well plates (Costar) were coated with 100 μl per wellof a 1:10 dilution of poly-L-lysine (Sigma) in PBS for 1 hour at roomtemperature. The plates were then washed twice with water, allowed toair-dry and placed at 4° C. overnight. One hundred μl of IM9 cells (at10⁶/ml in RPMI-1640) were then added to each well. Plates were thencentrifuged at 3200 rpm for 5 mins to pellet the cells. The media wascarefully aspirated and 200 μl of MPBS added to each well. The plateswere then left to block for 1 hour at room temperature.

To a separate 96-well plate, titrate test scFvs in MPBS, in triplicate,over a concentration range from 10 μg/ml down to 0.001 μg/ml were added.The final volume of test scFv in each well was 55 μl. Competition withunlabelled B Lymphocyte Stimulator was also included in every assay as acontrol. Unlabelled B Lymphocyte Stimulator, in MPBS, was typicallytitrated in triplicate, over a concentration range from 1 μg/ml down to0.001 μg/ml. 10 μl of biotinylated-B Lymphocyte Stimulator (at 162.5ng/ml) in MPBS was added to each well to give a final concentration of25 ng/ml. Plates were then incubated at room temperature for 30 minutes.

The IM9 coated plates was washed twice in PBS, tapped dry andimmediately 50 μl of the scFv/biotinylated-B Lymphocyte Stimulator mixwas added and incubated at room temperature for 1 hour. Plates werewashed three times in PBST and three times in PBS, tapped dry and 50 μlper well added of streptavidin-Delfia (Wallac) at 1:1000 dilution in theManufacturer's assay buffer. The plates were then incubated at roomtemperature for 1 hour and washed six times in Delfia wash solution(Wallac). After tapping the plates dry, 100 μl per well of Delfiaenhancement solution (Wallac) was added. The plates were gently tappedto encourage micelle formation, incubated at room temperature for 10minutes, and fluorescence read on a Wallac 1420 workstation at 6520 nM.

Typical titration curves for two scFv antibodies, I007F11 and I050A07,are shown in FIG. 6. Unlabelled B Lymphocyte Stimulator competed forbinding to its receptor with an IC₅₀ value of 0.8 nM. The IC₅₀ valuesfor I007F11 and I050A07 are 7.9 nM and 17.1 nM, respectively. The assaywas performed in triplicate and standard error bars are shown. The 9scFvs that demonstrated the greatest inhibition as scFv are listed inTable 4. This data also confirms that these9 scFvs recognize the solubleform of B Lymphocyte Stimulator.

TABLE 4 9 ScFvs that demonstrated greatest potency in B LymphocyteStimulator Receptor Binding Inhibition Assay ScFv Antibody I017D10I022D01 I008A11 I006D08 I031F02 I050A12 I050B11 1051C04 I003F12S

Example 8 Antibodies Recognizing a Soluble form of B LymphocyteStimulator

A library of phage was screened in an assay to identify those phagedisplaying scFvs that immunospecifically bind to the soluble but not themembrane-bound forms of B Lymphocyte Stimulator.

A phage library was screened for the ability to bind to biotinylated BLymphocyte Stimulator. The phage were exposed to biotinylated BLymphocyte Stimulator, allowed an interval of time to bind thebiotinylated B Lymphocyte Stimulator. Phage binding bio-B LymphocyteStimulator were then isolated by capture on streptavidin coated magneticbeads.

The phage identified in the screen above (capture of Bio-B LymphocyteStimulator from solution) were then screened by ELISA for their abilityto bind immobilized B Lymphocyte Stimulator. The scFv expressed by phagethat bound immobilized B Lymphocyte Stimulator were then cloned andsequenced. Again, several sequences were identified multiple times, thusa panel (panel 2) consisting of on example of each phage expressing aunique scFv was then characterized further.

The derived amino acid sequences of these scFvs are shown in Table 1above. The individual V_(H) and V_(L) segments of the scFvs were alignedto the known human germline sequences in V-BASE (Tomlinson et al, whichcan accessed on the United Kingdom Medical Research Council (MRC) Centrefor Protein Engineering website) and the closest germline identified.

Example 9 Specificity for Soluble B Lymphocyte Stimulator

The scFvs were isolated from a library of phage based on their abilityto bind a soluble form of B Lymphocyte Stimulator. Briefly, phage werepreincubated with biotinylated B Lymphocyte Stimulator in solution.Phage that bound to this biotinylated B Lymphocyte Stimulator were thenisolated using streptavidin coated magnetic beads.

The specificity of each of the unique scFvs for B Lymphocyte Stimulatorand for the membrane-bound form of B Lymphocyte Stimulator, wasdetermined by phage ELISA. B Lymphocyte Stimulator was immobilised ontoplastic as a purified soluble form of the protein or as a membrane-boundform present on plasma membrane preparations from the humanmacrophage-like cell line, U937. Maintenance of U937 cells and plasmamembrane preparations were performed as detailed in Example 2.

Phage ELISA

To determine the specificity of each of the scFvs, a phage ELISA wasperformed for each antibody against human B Lymphocyte Stimulator, U937plasma membranes, TNFα, BSA and an uncoated well. Antigen coatingconditions were as described in Example 2, apart from human B LymphocyteStimulator. B Lymphocyte Stimulator was first biotinylated (as describedin Example 3) and coated at 1 μg/ml onto streptavidin coated plates(Reacti-Bind, Pierce) for 30 mins at room temperature. The plates werethen washed, blocked and the phage ELISA performed as detailed inExample 2.

The results for 3 clones (I074B 12, I075F12 and I075A02) that bind thesoluble but not the membrane-bound form of B Lymphocyte Stimulator areshown in FIG. 7. As a control, a phage antibody that recognizes TNFα, isalso shown in FIG. 7. There is a small non-specific background signal onthe U937 plasma membranes that is evident with both the anti-BLymphocyte Stimulator scFvs as well as the anti-TNFα control. All 3anti-B Lymphocyte Stimulator scFvs recognize B Lymphocyte Stimulator butnot U937 plasma membranes, TNFα, BSA or an uncoated well (PBS only).This indicates that the scFvs do not bind the membrane-bound form of BLymphocyte Stimulator. Further, The fact that these scFvs were isolatedon the basis of their ability to bind soluble biotinylated B LymphocyteStimulator indicates that they bind the soluble form of B LymphocyteStimulator. Further confirmation of these scFvs' specificity for BLymphocyte Stimulator is provided in Example 10.

Example 10 Inhibition in an in vitro Receptor Binding Assay by PhagescFvs

All of the unique phage scFvs from panel 2 were assessed for theirability to inhibit B Lymphocyte Stimulator binding to its cognatereceptor on IM9 cells. The biotinylation of B Lymphocyte Stimulator,maintenance of IM9 cells and receptor binding inhibition assay wereperformed as described in Example 3.

Results for two phage scFvs, I0025B09 and I026C04 are shown in FIG. 8.Maximal binding of biotinylated B Lymphocyte Stimulator to its receptor(bio-B Lymphocyte Stimulator only), the background signal in the absenceof biotinylated B Lymphocyte Stimulator (no bio-B LymphocyteStimulator), and results with an irrelevant (i.e. does not recognize BLymphocyte Stimulator) phage antibody are also shown. Both phage scFvsinhibited biotinylated B Lymphocyte Stimulator binding to its receptoron IM9 cells. 33 of the unique scFvs from panel 2 were identified forfurther study. These 33 scFvs demonstrated the greatest inhibition asphage particles in this assay and are listed in Table 5.

TABLE 5 Identification of 33 phage scFvs to free B Lymphocyte Stimulatorthat demonstrate the most significant inhibition of biotinylated-BLymphocyte Stimulator binding to its receptor Antibody Antibody AntibodyAntibody I026C04 I074B12 I073F04 I065D04 I003C06 I075A02 I078D08 I068C08I025B09 I068B08 I078D02 I068F03 I027B12 I068B04 I075G01 I069B07 I025B06I068C06 I071B03 I030A10 I075F12 I072B09 I002A01R I065D08 I078H08I002A01K I065F08 I064C04 I026C04R I067B10 I064C07 I026C04K I067F05

Example 11 Specificity of anti-B Lymphocyte Stimulator scFvs

The specificity of the 33 scFvs (listed in Table 5) for immobilizedhuman and murine B Lymphocyte Stimulator was determined using phageELISA.

Phage ELISA

To determine the specificity of the 33 scFvs, a phage ELISA wasperformed as described in Example 4 against human and mouse B LymphocyteStimulator, and a panel of related human antigens: TRAIL, LIGHT, TNFα,TNFβ, and an uncoated well (PBS only).

Typical results for two scFvs, I067F05 and I078D02 are shown in FIG. 9.A control antibody that specifically recognizes TNFα is also shown. Bothanti-B Lymphocyte Stimulator scFvs specifically recognize immobilizedhuman and mouse B Lymphocyte Stimulator but not any other antigentested.

All 33 scFvs are specific for human B Lymphocyte Stimulator. 14/33cross-react with mouse B Lymphocyte Stimulator but not with any otherunrelated or related antigen tested.

Example 12 scFv Off-Rate Determinations

Off-rate determinations, preparation of a low density B LymphocyteStimulator surface and kinetic measurements were as detailed in Example6.

The binding curves for individual scFvs were analysed using theBIAevaluation software to determine antibody off-rates. Kinetic analysisfor a typical scFv antibody, I002A01, is shown in FIG. 10. I002A01 has aK_(off)=9×10⁻⁴ s⁻¹.

Example 13 Inhibition in an in vitro Receptor Binding Assay by scFvAntibodies

The 33 scFvs identified in Table 5 were prepared as purified scFvs andassessed for their ability to inhibit B Lymphocyte Stimulator binding toits receptor on IM9 cells. The scFvs were purified and analysed in thereceptor binding inhibition assay as described in Example 6.1.8.

Typical titration curves for two scFvs, I0068C06 and I074B12, are shownin FIG. 11. Unlabelled B Lymphocyte Stimulator competed for binding toits receptor with an inhibitory constant 50 (IC₅₀) value of 0.66 nM. TheIC₅₀ values for I0068C06 and I074B12 are 61 nM and 13 nM, respectively.The assay was performed in triplicate and standard error bars are shown.The 7 scFvs that demonstrated the greatest inhibition as scFv are listedin Table 6.

TABLE 6 Identification of 7 scFvs to free B Lymphocyte Stimulator thatdemonstrate the most significant inhibition of biotinylated-B LymphocyteStimulator binding to its receptor as purified scFv's. AntibodyI002A01-R I002A01-K I026C04-R I026C04-K I068C06 I075F12 I067B10

Example 14 ScFvs Recognizing Membrane-Bound B Lymphocyte Stimulator

A library of phage was screened in an assay to identify those phagedisplaying scFvs that immunospecifically bind to the membrane-bound butnot the soluble form of B Lymphocyte Stimulator.

As a starting point, a library of phage expressing scFv antibodies werepanned on immobilized HIS-tagged B Lymphocyte Stimulator. Phage isolatedby panning were then screened for the ability to bind to HIS-tagged BLymphocyte Stimulator. HIS-tagged B Lymphocyte Stimulator was obtainedby expressing amino acids 71-285 of SEQ ID NO:3228 using the pQE9 vector(Qiagen Inc., Valencia, Calif.) in E. coli and purifying the expressedprotein. This phage clones identified by this screen were thensequenced. After sequencing, A panel (panel 3) of phage each expressinga unique scFv that bound HIS-tagged B Lymphocyte Stimulator wasgenerated and further characterized.

The derived amino acid sequences of the unique scFvs from panel 3 areshown in Table 1 above. The individual V_(H) and V_(L) segments of thescFvs were aligned to the known human germline sequences in V-BASE(Tomlinson et al, which can accessed on the United Kingdom MedicalResearch Council (MRC) Centre for Protein Engineering website) and theclosest germline identified.

Example 15 Recognition of Membrane-Bound B Lymphocyte Stimulator

The specificity of each of the unique scFvs for both the membrane-boundform of B Lymphocyte Stimulator as well as for the soluble form of BLymphocyte Stimulator, was determined by phage ELISA.

B Lymphocyte Stimulator was immobilised onto plastic either directly asa purified soluble form of the protein or biotinylated and coated on astreptavidin plate as in Example 9. Binding to HIS-tagged B LymphocyteStimulator was used as a primary screen for scFv's that would bind themembrane-bound form of B Lymphocyte Stimulator (see below). Themembrane-bound form of B Lymphocyte Stimulator was presented as plasmamembranes preparations from the human macrophage-like cell line, U937 orthe murine cell line P388.

Mouse monoclonal antibodies have been raised against His-tagged BLymphocyte Stimulator according to standard procedures. Characterizationof these mouse monoclonal antibodies revealed that they specificallyrecognized both His-tagged B Lymphocyte Stimulator and themembrane-bound form of B Lymphocyte Stimulator on U937 cells, but notsoluble B Lymphocyte Stimulator. Therefore, specific recognition ofHis-tagged B Lymphocyte Stimulator was used as supporting evidence forthe recognition of the membrane-bound form of B Lymphocyte Stimulator byphage and scFv antibodies.

Phage ELISA

To determine the specificity of each of the scFvs, a phage ELISA wasperformed for each antibody against His-tagged human B LymphocyteStimulator, U937 plasma membranes, TNFα, BSA and an uncoated well.Antigen coating conditions were as described in 2. apart from human BLymphocyte Stimulator. B Lymphocyte Stimulator was first biotinylated(as described in Example 3) and coated at 1 μg/ml onto streptavidincoated plates (Reacti-Bind, Pierce) for 30 mins at room temperature. Theplates were then washed, blocked and the phage ELISA performed asdetailed in Example 2.

The results for 3 clones, I079C01, I081C10 and I082A02, and a controlphage antibody that recognizes TNFα, are shown in FIG. 12. All 3 scFvsrecognize U937 plasma membranes (U937) and His-tagged B LymphocyteStimulator (HIS-B Lymphocyte Stimulator) but not, biotinylated BLymphocyte Stimulator (bio-B Lymphocyte Stimulator) or an uncoated well(PBS). This indicates that the scFvs recognize the membrane-bound formof B Lymphocyte Stimulator.

Example 16 Specificity for Membrane-Bound B Lymphocyte Stimulator

The specificity of the scFvs for only the membrane-bound form of BLymphocyte Stimulator, and not for the soluble form, was confirmed usinga competition ELISA. This assay assesses the ability of test phage scFvsto bind to the membrane-bound form of B Lymphocyte Stimulator on U937plasma membranes in the presence of different forms of competing BLymphocyte Stimulator. Competing B Lymphocyte Stimulator was either theHis-tagged form of B Lymphocyte Stimulator or soluble B LymphocyteStimulator. ScFvs specific for the membrane-bound B LymphocyteStimulator would be expected to be competed out by pre-incubation withHis-tagged B Lymphocyte Stimulator but not by pre-incubation withsoluble B Lymphocyte Stimulator.

Maintenance of U937 cells and plasma membrane preparations wereperformed as detailed in Example 2.

Competition ELISA

U937 plasma membranes (50 μl per well) were coated at 10 μm/ml in PBSonto Falcon 96-well plates overnight at 4° C.

Individual E. coli colonies containing a phagemid representing one ofthe unique scFvs from the panel 3 were inoculated into 50 ml tubes(Falcon) containing 5 ml 2TYAG medium. Tubes were incubated at 37° C.for 4 hours, shaking. M13KO7 helper phage was added to each tube to anMOI of 10 and the tubes were incubated for a further 1 hour at 37° C.The tubes were centrifuged in a benchtop centrifuge at 3500 rpm for 10minutes. The supernatant was removed and cell pellets were resuspendedin 5 ml 2TYAK and incubated at 30° C. overnight, shaking. The next day,tubes were centrifuged at 3500 rpm for 10 min and the phage-containingsupernatant carefully transferred into a fresh tube.

For each test phage antibody, 3 aliquots of 20 μl 18% marvel/6×PBS weretransferred into separate wells of a 96-well plate. The first aliquotwas supplemented with His-tagged B Lymphocyte Stimulator to a finalconcentration of 60 μg/ml. The second aliquot was supplemented withsoluble B Lymphocyte Stimulator to a final concentration of 60 μg/ml.The third aliquot was not supplemented with any competing antigen. Onehundred μl of phage supernatant was then added to each aliquot and leftto block at room temperature for 1 hour.

The antigen-coated plates were washed once with PBS before the additionof 200 μl/well 3% marvel/PBS. These plates were left to block at 37° C.for 1 hour and were then washed once with PBS. Duplicate samples of 50μl pre-blocked phage (above) were added to the antigen-coated plates andleft at room temperature for 1 hour. Plates were washed 3× with PBS/0.1%Tween 20, then 3× with PBS. Fifty μl/well mouse anti-M13 HRP (Pharmacia)at 1/5000 in 3% Marvel/PBS was added and left for 1 hour at roomtemperature. Plates were washed 3 times with PBS/0.1% Tween 20, then 3times with PBS. Fifty μl/well HRP-labelled anti-mouse Envision polymer(DAKO) at 1/50 in 3% marvel/PBS was added and left for 1 hour at RT.Plates were washed 3 times with PBS/0.1% Tween 20, then 3 times withPBS. Next, 50 μl/well of TMB (Sigma) was added and plates left todevelop for 30 to 60 minutes. When sufficient color has developed, 25μ/well 0.5M H₂SO₄ was added to stop the reaction. The plates were readat 450 nm on a microtiter plate reader (Bio-Rad 3550).

The results for 3 clones, I079B04, I079F08 and I080B01, and a controlphage antibody that recognizes TNFα, are shown in FIG. 13. All 3 scFvsrecognize U937 plasma membranes (U937). This binding is competed out tobackground levels (i.e. comparable to the signal observed with theanti-TNFα phage antibody) in the presence of His-tagged B LymphocyteStimulator (HIS-B Lymphocyte Stimulator) but not biotinylated BLymphocyte Stimulator (bio-B Lymphocyte Stimulator). This confirms thatthe scFvs specifically recognize the membrane-bound form but not thesoluble form of B Lymphocyte Stimulator.

Example 17 High Throughput BIAcore Screen to Identify High AffinityscFvs

This is a 96-well screen where the test samples (scFvs) are derived from1 ml periplasmic extracts of individual antibody expressing clones.Potentially higher affinity scFvs are then identified principally asthose giving a large number of total RU's bound to a HIS-B LymphocyteStimulator surface in BIAcore. This method of ranking does assumeapproximately equal yields of scFv from each clone. Since this is notalways the case, some scFvs may also be identified that simply expresshigh levels of scFv. These can be discriminated from those of higheraffinity by further characterization of the scFvs (see Example 18).

Preparation of ScFv from 1 ml E. coli Cultures

Individual E. coli colonies containing a phagemid representing one ofthe unique scFvs from panel 3 were inoculated into 96-well platescontaining 100 μl 2TYAG medium per well. Eight wells on each plate werereserved for positive and negative control samples. The plate was grownovernight at 30° C. with shaking at 120 rpm.

Next day, 1 ml of 2TYAG+345 mM sucrose was added to each well of anautoclaved 96 deep well plate (Beckman). Twenty μl of each overnightculture was resuspended and transferred to the appropriate well of thedeep well plate. The plate was grown for approximately 3.5 hours at 30°C. with shaking at 250 rpm (or until the OD₆₀₀=0.6). Fifty μl of 1M IPTGwas added to 5 ml 2TY and 10 μl of this was added to each well. Theplate was grown overnight at 30° C. with shaking at 250 rpm.

Plates were kept at 4° C. for the remainder of the procedure. Theovernight plate (above) was centrifuged at 3500 rpm for 10 minutes at 4°C. to pellet the cells. The supernatant was decanted and each pelletresuspended in 100 μl TES (0.2M Tris HCl pH8.0, 0.5 mM EDTA, 0.5Msucrose) and transferred to a fresh 96 well plate. This plate wasincubated on ice for 30 minutes and then centrifuged for 10 minutes at3500 rpm at 4° C. to pellet the cell debris. During centrifugation, 15μl of freshly made protease inhibitors cocktail (Roche, 1 tabletdissolved in 1.5 ml water) was added to each well of a fresh 96 wellplate. Supernatants from the centrifuged plate were then transferred tothe plate containing the protease inhibitors. The plate was centrifugedat 3500 rpm for 10 minutes at 4° C. and the supernatant was transferredto a further 96-well plate. This step was repeated at least once more oruntil there was no sign of any cell debris following centrifugation.Finally, the plate was covered in foil to prevent evaporation of samplesduring the BIAcore run.

Generation of a High Density HIS-B Lymphocyte Stimulator Surface

All BIAcore analysis was performed on BIAcore 2000 machines, using theBIAcore 2000 control software and evaluated using the BIAevaluation 3.0software. A high density His-tagged B Lymphocyte Stimulator surface(>1000 RU HIS-B Lymphocyte Stimulator coupled) was prepared in flow cell2 by amine coupling to a CM5 chip. A new CM5 chip was inserted into theBIAcore and a sensorgram started over flow cell 2 with HBS buffer at aflow rate of 5 μl/min. The NHS and EDC solution were mixed 1:1 beforeinjecting 30 μl over the CM5 surface. Fifty μl HIS-B LymphocyteStimulator (at 10 μg/ml in Sodium acetate buffer, pH4) was injected andallowed to couple to the surface. Thirty μl of ethanolamine-HCl solutionwas then injected to block free NHS esters. Prior to using the chip, 10μl of 4M Guanidine hydrochloride in HBS was injected over the surface tostrip the surface of non-covalently bound B Lymphocyte Stimulator. Ablank surface (no HIS-B Lymphocyte Stimulator) was also prepared overflow cell 1 so that non-specific binding effects can be subtracted fromthe HIS-B Lymphocyte Stimulator binding curves.

Typically, a 5000 RU His-tagged B Lymphocyte Stimulator surface wasgenerated in this way and used for 96-well analysis of scFvs isolatedfrom the periplasm of E. coli.

BIAcore Analysis

The 96-well plate containing periplasmic scFvs was secured inside theBIAcore. Two ml of 4M Guanidine hydrochloride in HBS was placed in arack inside the BIAcore for regeneration of the HIS-B LymphocyteStimulator surface between samples. The sensorgram was run over flowcells 1 and 2 at a flow rate of 200 minute. The following method wasrun:

MAIN FLOWCELL 1,2,3,4 LOOP cycle STEP APROG inj %pos ENDLOOP APPENDCONTINUE END DEFINE LOOP cycle LPARAM %pos r1a1 r1b1 r1c1 r1d1 r1e1 r1f1etc (all wells liste until r1h12) END DEFINE APROG inj PARAM %pos FLOW20 KINJECT %pos 35 30 !scfv injection QUICKINJECT r2f3 10 !regenerationEXTRACLEAN END

When the run had finished, the sensorgram data for flow cell 1 wassubtracted from the data for flow cell 2 for each sample using theBIAevaluation software. The clones were compared with one anotherprincipally by overall RU change as the scFv dissociates from thesurface. In addition a few scFvs were identified as having potentiallyslower off-rates. An example of the dissociation section of a typicalsensorgram for 8 scFvs is shown in FIG. 14. An anti-TNFα antibody thatdoes not recognize B Lymphocyte Stimulator was included as a control. Ofthe 8 scFvs exemplified, I079F06 was identified for further study due tothe relatively high numbers of RU's bound to the surface.

ScFvs were identified principally if they demonstrated a RU change ofover 1200, a few were also identified as having potentially slower thantypical off-rates. A total of 28 clones were chosen on these criteriaand are listed in Table 7.

TABLE 7 Identification of 28 antibodies to membrane- bound B LymphocyteStimulator that demonstrate the most significant RU changes by BIAcoreAntibody Antibody I079C01 I084C04 I082H08 I080E05 I079E02 I083B12I079B05 I082G01 I079F06 I082G02 I079F08 I082C03 I079F11 I082A05 I079B12I082D07 I080B01 I082B08 I080G09 I084A01 I099D03 I084B02 I080D03 I080A08I080A03 I084C11 I083G03 I080G07

Example 18 scFv Affinity Determinations

The affinity (K_(D)) of the 28 scFvs was determined using the BIAcore.

Low Density HIS-B Lymphocyte Stimulator Surface for Kinetic Studies

500RU surfaces were used for kinetic studies of purified scFv binding toHIS-B Lymphocyte Stimulator. The method to prepare these surfaces wasidentical to the method described in Example 17, only smaller volumes ofHIS-B Lymphocyte Stimulator were injected.

Measurement of scFv Binding Kinetics

The chip containing the low density HIS-B Lymphocyte Stimulator surfacewas inserted into the BIAcore. A dilution series for each of the 28purified scFvs (prepared as in Example 6) were diluted in HBS (typicallystarting with 50 μg/ml scFv and double diluting down to 1.5 μm/ml). Thedilution series was then injected sequentially over the blank control(flow cell 1) and low density HIS-B Lymphocyte Stimulator surface (flowcell 2) using the following program:

MAIN FLOWCELL 1,2,3,4 APROG genab r1d1 ab1 APROG genab r1d2 ab2 APROGgenab r1d3 ab3 APROG genab r1d4 ab4 APROG genab r1d5 ab5 APROG genabr1d6 ab6 APPEND CONTINUE END DEFINE APROG genab PARAM %Abpos %AbId FLOW20 KINJECT %Abpos 200 80 INJECT r2f3 10 EXTRACLEAN END

Bound scFv were removed by injecting 10 μl of 4M Guanidine hydrochloridein HBS (location r2f3 in the above program) over the surface betweensamples. Binding curves for individual scFv were analysed using theBIAevaluation software to determine antibody on- and off-rates.

A typical example of the binding curves generated for the scFv antibodyI082C03 is shown in FIG. 15. The off-rate for this clone was calculatedas 2×10⁻³ s⁻¹. The affinity of I082C03 was calculated as 20 nM, assuming100% activity of the scFv. The 5 scFvs with the highest affinities asscFvs are given in Table 8.

TABLE 8 Identification of 5 antibodies to membrane-bound B LymphocyteStimulator that have the highest affinities as scFvs Antibody Affinity(K_(D)) I079F11 5 nM I079E02 10 nM  I082G02 6 nM I082H08 1 nM I099D03 4nM

Example 19 Recognition of Mouse Membrane-Bound B Lymphocyte Stimulator

The ability of the 5 scFvs listed in Table 8 to also recognize murinemembrane-bound B Lymphocyte Stimulator was determined using acompetition ELISA. This assay assesses the ability of test phage scFvsto bind to the membrane-bound form of B Lymphocyte Stimulator on themurine cell line, P388, plasma membranes in the presence of differentforms of competing human B Lymphocyte Stimulator. Competing B LymphocyteStimulator was either presented as the His-tagged form of B LymphocyteStimulator, or soluble B Lymphocyte Stimulator. ScFvs that recognizemouse membrane-bound B Lymphocyte Stimulator would give an ELISA signalon the P388 plasma membranes that is competed out by pre-incubation withHIS-tagged B Lymphocyte Stimulator but not by pre-incubation withsoluble B Lymphocyte Stimulator.

Maintenance of P388.D1 Cells and Preparation of Plasma Membranes

P388.D1 cells are a mouse monocyte-macrophage like cell line. They werecultured in L-15 medium supplemented with 2 mM L-glutamine, 10% CS, 10Upenicillin, 100 g/ml streptomycin (all reagents from Sigma). Cells weresplit 1:4 every 3-4 days to maintain a cell density of 2-8×10⁵ per ml. Afresh aliquot of cells was thawed from liquid nitrogen every 6 weeks.Plasma membrane fractions were prepared as described in Example 2.

Competition ELISA

P388 plasma membranes (50 μl per well) were coated at 10 μg/ml in PBSonto Falcon 96-well plates overnight at 4° C. The method is otherwiseessentially as described Example 16.

The results for 3 clones, I079E02, I082H08 and I099D03 are shown in FIG.16. All 3 scFvs recognize P388 plasma membranes. This binding iscompeted out in the presence of HIS-tagged B Lymphocyte Stimulator(HIS-B Lymphocyte Stimulator) but not in the presence of biotinylated BLymphocyte Stimulator (bio-B Lymphocyte Stimulator). This confirms thatthese scFvs also recognize the membrane-bound form but not the solubleform of mouse B Lymphocyte Stimulator.

Example 20 Conversion of scFvs to IgG1 Format

The VH domain and the VL domains of scFvs that we wished to convert intoIgG molecules were cloned into vectors containing the nucleotidesequences of the appropriate heavy (human IgG1) or light chain (humankappa or human lambda) constant regions such that a complete heavy orlight chain molecule could be expressed from these vectors whentransfected into an appropriate host cell. Further, when cloned heavyand light chains are both expressed in one cell line (from either one ortwo vectors), they can assemble into a complete functional antibodymolecule that is secreted into the cell culture medium. Methods forconverting scFvs into conventional antibody molecules are well knownwithin the art.

Generation of NS0 Cell Lines Expressing Anti-B Lymphocyte StimulatorAntibodies (IgG1)

Plasmids containing the heavy and light chains were separatelylinearized using the Pvu I restriction enzyme. The linearized DNAs werepurified by phenol-chloroform extraction followed by ethanolprecipitation and then resuspended in H₂O. NS0 cells (10⁷) from agrowing culture were electroporated (0.25 kV and 975 μF) in PBS with12.5 μg linearized heavy chain plasmid DNA and 37.5 μg linearized lightchain DNA. The cells were washed in 20 ml non-selective medium (10% FCSin DMEM supplemented with 6 mM glutamine, amino acids andpenicillin/streptomycin) and then transferred in 12.5 ml medium into aT75 cm² flask and incubated overnight at 37° C., 5% CO₂/air. The dayafter transfection the cells were resuspended in selective mediumcontaining 1 mg/ml geneticin and dispensed into 5×96-well plates at 200μl/well. After 18 days at 37° C. (5% CO₂/air) the colony supernatantswere screened by an ELISA that detects assembled human IgG in order toidentify colonies expressing IgG. Approximately twenty positive colonieswere expanded and adapted to growth in serum-free, selective medium.Duplicate T25 cm² flasks were set up. Cells from one flask were frozendown as a stock and cells in the second flask were grown to saturation.The productivity of the saturated cultures was assessed by ELISA. Thehighest producing cell lines were then selected for large-scale antibodyproduction.

The above procedure is exemplified for the I006D08 anti-B LymphocyteStimulator antibody constructs. Following electroporation and selectionof NS0 cells, supernatants from ninety-three wells each containing asingle colony were screened by ELISA to detect assembled IgG1, antibody.Twenty-seven of the supernatants were identified as containing IgG. Thecolonies from 24 of the positive wells were transferred to 1 mlselective medium in a 24-well plate and allowed to grow for 2 days. The1 ml cultures of cells were then added to 4 ml selective mediumcontaining reduced serum (0.5% FCS) in a T25 cm² flask. When thecultures reached confluency 1 ml cells were diluted in 4 ml selective,serum-free medium in a T25 cm² flask. At confluency this subcultureregime was repeated again. Finally 1 ml cells from the culturecontaining 0.1% FCS was diluted with 9 ml serum-free, selective mediumand divided into 2×T25 cm² to form the saturated and stock cultures. Thestock cultures were frozen down and stored in liquid nitrogen once thecultures were confluent. The saturation culture was grown until theviability of the culture was <10%. Twenty-three out of the 24 coloniesoriginally expanded were successfully adapted to growth in serum-freemedium. The productivity of these serum-free adapted cell lines rangedfrom 0.3 to 17 μg/ml by ELISA quantification of the saturated, 5 mlserum-free cultures. The I006D08-32 cell line produced 17 μg/ml.

Large-Scale IgG Production

The highest-producing cell lines were revived from frozen stocks andthen expanded to 400 ml in selective, serum-free medium in 2 literroller bottles. The cells were grown at 37° C. and rolled at 4 rpm withthe headspace being re-equilibrated with 5% CO₂/air every 2-3 days.Finally the culture was expanded to a 4 liter volume by the addition ofserum-free medium without selection (400 ml per 2 liter roller bottle).The cultures were then grown to saturation.

This procedure is exemplified by the production of I006D08 antibody fromthe I006D08-32 cell line. The frozen stock of I006D08-32 was revivedinto a T25 cm² containing 5 ml serum-free medium containing 1 mg/mlgeneticin and grown at 37° C. in 5% CO₂/air incubator. After two daysgrowth the culture was diluted with 7.5 ml fresh medium and transferredto a T75 cm² flask. After a further three days in the incubator thecells were transferred to 130 ml selective medium and transferred to a 2liter roller bottle. After three days growth the cells were diluted with500 ml selective medium and split into 2×2 liter roller bottles. Afteranother 2 days 100 ml fresh selective medium was added to each roller.Finally the next day the culture was expanded to a total volume of 4liters with non-selective medium and divided into 10×2 liter rollerbottles. After three days the medium was supplemented with 6 mMglutamine. The cells were grown for 17 days from the final subcultureinto a 4 liter volume. The cells grew up to 3×10⁶ cells/ml beforeviability declined to <0.2×10⁶ cells/ml. At this low viability theculture supernatants were harvested. ELISA analysis indicated that theculture supernatant contained 33 μg/ml IgG. Hence, the 4 liter culturecontained 132 mg IgG.

IgG Purification

The purification of the IgG from the fermentation broth is performedusing a combination of conventional techniques commonly used forantibody production. Typically the culture harvest is clarified toremove cells and cellular debris prior to starting the purificationscheme. This would normally be achieved using either centrifugation orfiltration of the harvest. Following clarification, the antibody wouldtypically be captured and significantly purified using affinitychromatography on Protein A Sepharose. The antibody is bound to ProteinA Sepharose at basic pH and, following washing of the matrix, is elutedby a reduction of the pH. Further purification of the antibody is thenachieved by gel filtration. As well as removing components withdifferent molecular weights from the antibody this step can also be usedto buffer exchange into the desired final formulation buffer.

Purification of I006D08 IgG1

The harvest was clarified by sequential filtration through 0.5 μm and0.22 μm filters. Clarified harvest was then applied to a column ofrecombinant Protein A Sepharose equilibrated at pH 8.0 and washed withthe equilibration buffer. I006D08 antibody was eluted from the Protein ASepharose by application of a buffer at pH3.5. The collected antibodycontaining eluate was then neutralized to pH 7.4 by the addition of pH8.0 buffer. The neutralized eluate was concentrated by ultrafiltrationusing a 30 KDa cut off membrane. Concentrated material was then purifiedby Sephacryl S300HR gel filtration using phosphate buffered saline asthe mobile phase. The final monomeric IgG1 fraction from the gelfiltration column was then concentrated to the desired formulationconcentration by ultrafiltration using a 30 KDa cut off membrane. Thefinal product was filtered through a 0.22 μm filter.

Example 21 Antibody Neutralization of Murine Splenocyte Proliferation asMeasured by 3HdT Incorporation

To determine if an antibody inhibited B Lymphocyte Stimulator mediated Bcell proliferation, a splenocyte proliferation assay was performedBriefly, murine splenocytes were isolated by flushing spleen using a 25g needle and 10 ml of complete medium (RPMI 1640 with 10% FBS containing100U/ml penicillin, 100 μg/ml streptomycin, 4 mM glutamine, 5×10⁻⁵Mβ-mercaptoethanol). The cells were passed through a 100 micron nylonfilter to remove cell clumps. The cell suspension was then ficolled at400×g for 25 minutes at room temperature (one 15 ml conical tube/spleen;3 ml ficol, 10 ml cell suspension/spleen; Ficol 1083 from Sigma). Therecovered cells were washed 3 times in complete medium and counted.Recovered cells were then diluted to a concentration of 3×10⁶/ml incomplete medium containing a 3× concentration of SAC (3×=1:33,333dilution of stock; stock is a 10% suspension of Staph. aureus (Cowan Istrain) available from Calbiochem).

For each antibody, 50 microliters of antibody dilutions at 30 μg/ml, 3.0μg/ml, and 0.3 μg/ml concentrations were aliquotted into individualwells of a 96 well plate in triplicate. Suitable positive controls, suchas, for example monoclonal antibody 15C10, were also used. Antibody15C10 is described in Moore et al., (1999), Science 285: 260-263 andWO0050597; 15C10 has also been deposited with the ATCC™ on Feb. 1, 2000as ATCC™ Deposit No. PTA-1158. Medium containing no antibody (and humanisotype controls (purchased commercially) when necessary) were used asnegative controls.

B Lymphocyte Stimulator protein was diluted in complete medium toconcentrations of 300 ng/ml, 90 ng/ml and 30 ng/ml. 50 microliters ofeach of the B Lymphocyte Stimulator dilutions were then added to theantibody dilution series in the plates. The plate containing theantibody and B Lymphocyte Stimulator dilutions are then incubated for 30minutes at 37° C., 5% CO₂, after which 50 microliters of the splenocytecell suspension containing SAC was added to all wells. The plates werethen incubated for 72 hours (37° C., 5% CO₂).

After 72 hours, each well was supplemented with 50 μl of complete mediumcontaining 0.5 μCi of 3H-thymidine (6.7 Ci/mM; Amersham) and cells wereincubated for an additional 20-24 hours at (37° C., 5% CO₂). Followingincubation cells were harvested using a Tomtec Cell Harvester andfilters counted in a TopCount Scintillation counter (Packard).

Example 22 Human B Cell Proliferation Assay for in vitro Screening of BLymphocyte Stimulator Antagonist Molecules

The bioassay for assessing the effects of putative B LymphocyteStimulator antagonists was performed in triplicate in 96 well format bymixing equal volumes of B Lymphocyte Stimulator, responder cells, andputative antagonist each of which is prepared as a 3× stock reagent.

B-lymphocytes were purified from human tonsil by MACS (anti-CD3depletion), washed, and resuspended in complete medium (CM) (RPMI 1640with 10% FBS containing 100U/ml penicillin, 100 μg/ml streptomycin, 4 mMglutamine, 5×10E-5 M beta-mercaptoethanol) at a concentration of 3×10e6cells/mL. Staphylococcus aureus, Cowan I (SAC, CalBiochem) was added tocells at 3× concentration (3×=1:33,333 dilution of stock

Meanwhile, eight serial dilutions (3-fold) of potential antagonist wereprepared in CM such that the diluted antagonists are at 3× the finalconcentrations to be tested in the assay. Antibodies are routinelytested starting at a final concentration of 10 ug/mL and going down toabout 1.5 ng/mL.

Human rB Lymphocyte Stimulator was prepared in CM to 3× concentration(3×=300 ng/mL, 30 ng/mL, and 3 ng/mL) in CM. Potential inhibitors wereroutinely tested at several concentrations of B Lymphocyte Stimulator toavoid false negatives due to unexpectedly low affinity or antagonistconcentration.

Fifty microliters of diluted antagonist and 50 uL of diluted BLymphocyte Stimulator were added to the putative antagonist dilutionseries.

Cells were then incubated for 72 hours (37° C., 5% CO₂) in a fullyhumidified chamber. After 72 hrs., the cells were supplemented with 0.5μCi/well 3H-thymidine (6.7 Ci/mmol) and incubated for an additional 24hours. Plates were harvested using a Tomtec Cell Harvester and filterscounted in a TopCount Scintillation counter (Packard).

Example 23 Characterization of I006D08 and I116A01

I116A01 (SEQ ID NO:327) is a CDR3 mutant of the I006D08 (SEQ ID NO:2).The I116A01 scFv was subsequently isolated from a single chain phagedisplay library containing a repertoire of I006D08 mutations generatedby randomization of the last 6 amino acids of the heavy chain CDR3region of I006D08. The I116A01 scFv contains 7 amino acid differencesfrom the parental I006D08 sequence, 1 change in the VH framework 3region, 5 changes in the VH CDR3, and 1 change in the VLCDR3. Each ofthe assays described herein were performed using whole IgG1 moleculescomprising the VH and VL regions of I006D08 and I116A01.

Binding Studies of I006D08 and I116A01

Equilibrium dissociation constants (Kd) for I116A01 and I006D08 bindingto B Lymphocyte Stimulator were assessed using BIAcore technology(BIAcore, Uppsala, Sweden). Briefly, antibodies were captured onlow-density Protein A surfaces and B Lymphocyte Stimulator was passedover at various flow rates. The binding of B Lymphocyte Stimulator toimmobilized antibody was detected by changes in surface charge that wereproportional to the amount of bound B Lymphocyte Stimulator. Bindingvalues were calculated using software provided by the manufacturer.

I116A01 was determined to have an equilibrium dissociation constant, Kd,of 267±70 pM (mean±SD, n=3). The Kd value of I006D08, the parentalantibody, was determined to be 498±151 pM (mean±SD, n=3). This wasapproximately 2-fold higher than the Kd obtained for I116A01. Thedifference in affinity can be attributed to a 2-fold higher off rate(kd, 1/s) of I006D08 compared with I116A01, the on rates (ka, 1/Ms)being essentially equal (Table 9).

TABLE 9 Kinetic parameters for I116A01 and I006D08 binding to BLymphocyte Stimulator ka, 1/Ms kd, 1/s Kd, pM Mab Mean ± SD Mean ± SDMean ± SD I116A01 1.26 ± 0.14 × 10⁶ 3.32 ± 0.63 × 10⁻⁴ 267 ± 70 I006D081.43 ± 0.13 × 10⁶  5.8 ± 0.24 × 10⁻⁴ 498 ± 151

The Kd values obtained for the B Lymphocyte Stimulator antibodiesindicate that these antibodies bind to B Lymphocyte Stimulator with highaffinity and possess a slow dissociation or off rate.

The binding characteristics of I116A01 and I006D08 were investigatedfurther using a competitive binding ELISA in which B LymphocyteStimulator was immobilized directly on the surface of the well tocompare epitopes. Biotinylated I006D08 binding was inhibited in adose-dependent manner by the addition of unlabeled I006D08 or I116A01suggesting that these antibodies recognize overlapping or identicalepitopes on B Lymphocyte Stimulator.

Ability of I006D08 and I116A01 to Inhibit B LymphocyteStimulator-Induced Splenocyte Proliferation

The ability of I006D08 and I116A01 to inhibit B LymphocyteStimulator-induced Splenocyte proliferation was tested using the assaydescribed in Example 21 with slight modifications. In particular, theassay was modified such that the final concentration of B LymphocyteStimulator in the assay was 3ng/ml and the final dilution factor of SACwas 1:100,000. Using this assay it was shown that both antibodiesI006D08 and I116A01 were each found to inhibit murine splenocyteproliferation induced by both human (aa134-285 of SEQ ID NO:2) andmurine soluble B Lymphocyte Stimulator. The IC₅₀ values for theinhibition of human B Lymphocyte Stimulator induced-proliferation ofmurine B cells was 0.09 nM±0.01 nM and 0.06 nM±0.01 nM, respectively.

Additional Characterization of I006D08 and I116A01

I006D08 and I116A01 were identified herein as antibodies that bind boththe membrane bound and soluble forms of B Lymphocyte Stimulator. Theability to bind the membrane bound form of B Lymphocyte Stimulator wasdetermined according to the assays described above, e.g., ability tobind HIS-tagged B Lymphocyte Stimulator and ability to bind U937 and/orP388 membrane preparations in an ELISA assay. Further characterizationof antibodies I006D08 and I116A01 reveals that these antibodies bindrecombinant B Lymphocyte Stimulator polypeptides expressed on thesurface of a 293T cell. On the other hand, these antibodies are not ableto bind naturally occurring B Lymphocyte Stimulator (non-recombinant)expressed on the surface of a cell, e.g., K-562 cells (ATCC™ #CCL-243)or U-937 cells (ATCC™ #CRL-1593.2). Thus, I006D08 and I116A01 bind thesoluble form of B Lymphocyte Stimulator. I006D08 and I116A01 also bindthe membrane bound form of B Lymphocyte Stimulator (as determined bytheir ability to bind HIS-tagged B Lymphocyte Stimulator and theirability to bind U937 and/or P388 membrane preparations in an ELISAassay). I006D08 and I116A01 also bind recombinant B LymphocyteStimulator expressed on the surface of a 293T transfected fibroblastcell line, but do not bind naturally occurring B Lymphocyte Stimulatorprotein on the surface of a K-562 or U-937 cell.

Example 24 In Vivo Studies of Anti-B Lymphocyte Stimulator antibodies

Overexpression of B Lymphocyte Stimulator in transgenic animals resultsin an autoimmune phenotype including increased numbers of mature B cells(leading to enlarged spleens), increased levels of serumimmunoglobulins, as well as increased levels of autoantibodies (e.g.,anti-dsDNA antibodies) (McKay et al., (1999), The Journal ofExperimental Medicine 190:1697-1710 and Gross et al., (2000) Nature404:995-999, both of which are hereby incorporated by reference in theirentireties.) Similar effects are observed when recombinant B LymphocyteStimulator is administered to animals such as mice and monkeys. Theability of anti-B Lymphocyte Stimulator antibodies of the invention toreduce or prevent these B Lymphocyte Stimulator induced effects may betested through either co-administration or sequential administration ofsoluble B Lymphocyte Stimulator and anti-B Lymphocyte Stimulatorantibodies of the invention in an animal. An anti-B LymphocyteStimulator antibody of the invention would be considered neutralizing ifit prevented or reduced B Lymphocyte Stimulator-induced increases in Bcell numbers (resulting in lack of, or less, enlargement of spleen), andserum immunogloblins, and in particular serum autoantibodies.

Example 25 Antibody Production and Purification

The following example describes a large scale antibody production andpurification methods that can be used to make antibodies of the presentinvention. One of skill in the art will be aware of routinemodifications to the protocol described below, for example, as regardscolumn choice, column, loading, wash, and elution buffers, and pH.

Cell Culture Scale-Up and Antibody Production

A serum-free and animal source-free growth medium is used from thawingcells through scale-up to the production bioreactor. The growth mediumis prepared by adding 1 kilogram AGT powder granules preformulated withGS supplement to water for injection (WFI) quality water making a totalof 51 kilograms CD hybridoma medium. To the CD hyrbridoma medium, 1mL/kg cholesterol (synthetic) lipid concentrate (1000×). The AGT powdergranules preformulated with GS supplement for preparing the CD hybridomamedium and the synthetic cholesterol lipid concentrate and are availablefrom Invitrogen Corp., Carlsbad, Calif. The medium, hereinafter referredto as INV-CDH medium, is stored at 2-8° C. until use.

Thawing Cells from into Spinner Flask

Approximately 48×10⁶ NS0 cells engineered to express an antibody of theinvention, preferably in whole IgG1 format, are thawed at 37° C. in awater bath. The cells are transferred into a 250 mL spinner flask toyield approximately a 150 mL working volume with an inoculation densityof approximately 3.2×10⁵ cells/mL. The spinner flask(s) is then placedon magnetic stirrers in a humidified CO₂ incubator at 37° C. with 5% CO₂for 4 days. The agitation rate for the spinner flask is 80 rpm.

First Expansion(s) of Culture in Spinner Flask

The culture is aseptically expanded into a 1000 mL spinner flask to giveapproximately 600 mL working volume, at an inoculation cell density ofapproximately 2.5×10⁵ cells/mL. The spinner flask is then placed onmagnetic stirrers in a humidified CO₂ incubator at 37° C. with 5% CO₂for 3 days. The agitation rate for the spinner flask is 80 rpm.

The culture is again expanded aseptically into two 3000 mL spinnerflasks to give approximately 2×2000 mL working volume, at an inoculationcell density of approximately 3.0×10⁵ cells/mL. The spinner flask isthen placed on magnetic stirrers in a humidified CO₂ incubator at 37° C.with 5% CO₂ for 3 days. The agitation rate for the spinner flasks is 80rpm. At the same time, a reserve culture is created in a fresh 250 mLspinner flask. The reserve culture may be used as the seed for anotherlot as needed.

Seed Culture (25 Liters)

The 25 liter seed bioreactor is equipped with one impeller for mixing, adissolved oxygen probe, a temperature probe, a pH probe, asepticsampling and additional systems. The first step of the cell cultivationprocess is the addition of INV-CDH medium into the bioreactor. After themedium temperature reaches 37±0.5° C., the dissolved oxygen (DO) and pHlevels are stabilized by addition of N₂ and CO₂ to decrease dissolvedoxygen concentration to 25 to 75% air saturation, and obtain a pH of7.20±0.2. The agitation rate is 70 rpm. The pooled cell culture istransferred aseptically to the 25 liter seed bioreactor containing 21liters of sterile INV-CDH medium. During the cultivation process thetemperature is maintained via a plant stem and chilled glycol, whichregulate the temperature of the circulating jacket. The oxygenconcentration is maintained via oxygen and nitrogen sparging and surfaceaeration, and pH is controlled by addition of CO₂ gas to lower the pH.The cultivation period is 4 days. The bioreactor air/gas inlets and ventlines are protected by hydrophobic 0.2 μm filters.

Seed Culture (200 Liters)

The 200 L seed bioreactor is equipped with 1 impeller for mixing, adissolved oxygen probe, a temperature probe, a pH probe, asepticsampling and additional systems. The first step of the cell cultivationprocess is the addition of INV-CDH medium into the bioreactor. After themedium temperature reaches 37±0.5° C., the DO and pH levels arestabilized by addition of N₂ and CO₂ to decrease dissolved oxygenconcentration to 25 to 75% air saturation, and obtain a pH of 7.20±0.2.The agitation rate is 35 rpm. The 25 L seed culture is transferredaseptically to the 200 L seed bioreactor containing 175 L sterilemedium. During the cultivation process the temperature is maintained viaplant steam and chilled glycol which regulate the temperature of thecirculating jacket. The oxygen concentration is maintained via oxygenand nitrogen sparging and surface aeration, and pH is controlled byaddition of CO₂ gas to lower the pH. The cultivation period is 4 days.The bioreactor air/gas inlet and vent lines are protected by hydrophobic0.2 μm filters.

Production Culture (1600 Liters)

The production bioreactor is equipped with 2 impellers for mixing, adissolved oxygen probe, a temperature probe, a pH probe, asepticsampling and additional systems. 600 L of INV-CDH medium is asepticallytransferred into the 1600 L production bioreactor. After the mediumtemperature reaches 37±0.5° C., the DO and pH levels are stabilized byaddition of N₂ and CO₂ to decrease dissolved oxygen concentration to 25to 75% air saturation, and obtain a pH of 7.20±0.2. The agitation rateis 17 rpm. The 200 L seed culture is aseptically transferred into theproduction bioreactor. During the cultivation process the temperature ismaintained via plant steam and chilled glycol which regulate thetemperature of the circulating jacket, the oxygen concentration ismaintained via oxygen and nitrogen sparging and surface aeration, and pHis controlled by addition of CO₂ gas to lower the pH. The bioreactorair/gas inlet and vent lines are protected by hydrophobic 0.2 μmfilters. After 48 hours, 800 Liters of INV-CDH medium is transferredinto the production bioreactor in order to obtain the final volume. Theculture is cultivated in the production bioreactor 43 additional days.

Recovery and Purification Harvest of Cell Supernatant

Cell supernatant, (e.g., culture supernatant from NSO cells expressingantibodies of the invention) is harvested on day 5 or 6 post finalfeeding in the final production bioreactor. Cell culture brothtemperature is cooled down to 15° C. in the recovery vessel at the timeof harvest and maintained at that temperature during the recovery.Centrifugation followed by depth filtration is used for cell removal andantibody recovery. The filtration process train consists of 0.1 μmnominal rated depth filter and 0.2 μm membrane filters connected inseries. A constant flow rate of 13±1 L/min is maintained during theoperation. The 0.2 μm filtered culture supernatant is collected in aprocess vessel, warned to 20° C. and and transferred to purification.The purification process is conducted at 22 to 26° C.

Chromatography on rProtein A Sepharose FF Column

The culture supernatant is loaded onto rProtein-A Sepharose FF column(affinity chromatography, Amersham Pharmacia) or equivalent column thatis equilibrated in 50 mM Tris-HCl, 0.5 M NaCl, pH 7.5. The rProtein-ASepharose FF column is washed with 0.05 M sodium citrate, 0.15 M sodiumchloride, pH 5.2 and eluted with 0.05 M sodium citrate, 0.15 M sodiumchloride, pH 3.2. The elution is monitored by ultraviolet (UV)absorbance at 280 nm. The elution peak is collected, and analyzed byA₂₈₀ and SDS-PAGE.

Virus Inactivation

The eluate from the rProtein-A Sepharose FF column is adjusted with 1 Mcitric acid to pH 3.4±0.2 and allowed to stand for 45-60 minutes forviral inactivation. The solution is then readjusted to pH 5.4 with 1 MTris base. The pH adjusted material is 0.2 μm filtered and stored at 20°C. up to 20 hours.

Chromatography on SP Sepharose FF Column

The inactivated material from the rProtein-A Sepharose FF column isdiluted with water for injection (WFI) to a conductivity of 7 mS, andloaded onto a SP Sepharose FF column (cation exchanger,Amersham-Pharmacia), or equivalent column equilibrated with 50 mM sodiumacetate, 30 mM sodium chloride, pH 5.4. The antibody is eluted from theSP column with 50 mM sodium acetate, 100 mM sodium chloride, pH 6.0. Theelution is monitored by ultraviolet (UV) absorbance at 280 nm. Theelution peak is collected and analyzed by A₂₈₀ and SDS-PAGE.

Virus Removal Filtration, Diafiltration and Concentration

The eluate from the SP Sepharose FF column is filtered through asequentially connected 0.2 μm filter and a virus removal filter (PallDV50 Ultipor® VF Filter System). The DV50 filtrate is stored at 20° C.up to 8 hours.

The DV50 filtrate is placed into a 30 kD molecular weight cut-offmembrane device (Millipore Pellicon) to concentrate to a targetconcentration of 35-40 mg/mL. The concentrated material is diafilteredagainst 10 mM sodium citrate, 1.9% glycine, 0.5% sucrose, pH 6.5. Thediafiltered material is monitored by A₂₈₀. The diafiltered bulk is 0.2μm filtered and stored at 20° C. up to 8 hours.

Chromatography on Q Sepharose FF Column

The diafiltered antibody solution is passed over a Q Sepharose FF column(anion exchanger, Amersham-Pharmacia) or equivalent column equilibratedwith 10 mM sodium citrate, 1.9% glycine, 0.5% sucrose, pH 6.5. Theantibody is collected in the flow-through and monitored by A₂₈₀. Theelution peak is collected and analyzed by A₂₈₀

Bulk Formulation, Filtration and Bulk Drug Substance Fill

Polysorbate 80 (1% stock solution in 10 mM sodium citrate, 1.9% glycineand 0.5% sucrose) is pre-filtered through a 0.2 μm filter and added tothe antibody solution to a final concentration of 0.01%. The purifiedantibody is aseptically filtered under a laminar flow hood through a 0.2μm filter and filled into polypropylene containers.

Storage of Bulk Drug Substance

The bulk drug substance is stored at 2-8° C. (short-term storage) or ator below −65° C. (long-term storage).

In Process Testing

In-process testing of bioreactor culture at harvest for each batch andin-process testing during the purification process are performed. Thebioreactor is sampled aseptically and the culture is tested at varioustimes throughout cultivation for cell density, viability and nutrientdetermination to ensure consistency of material being supplied forpurification. The purification process is monitored at each step.Appearance is checked by visual inspection. The protein concentration isdetermined by Absorbance at 280 nm. The pH of the material is checked.Purity is checked, for example, by SDS-PAGE and size exclusionchromatography. An ELISA may be performed to check the ability of theantibody to bind its antigen. The biological activity of the antibody isalso monitored. Residual DNA content, Endotoxin levels, and thebioburden (the number of viable organisms present in the antibodypreparation) are all monitored and kept at or below standard acceptablelevels. Additionally the oligosaccharide content may be analyzed; thepeptide sequence of the antibody chains may also be analyzed usingN-terminal sequencing and peptide mapping. Short and long-term studiesof antibody stability may also be performed.

It will be clear that the invention may be practiced otherwise than asparticularly described in the foregoing description and examples.Numerous modifications and variations of the present invention arepossible in light of the above teachings and, therefore, are within thescope of the appended claims.

The entire disclosure of each document cited (including patents, patentapplications, journal articles, abstracts, laboratory manuals, books, orother disclosures) in this application is incorporated in theirentireties herein by reference. Further, the sequences disclosed hereinare also disclosed in the Sequence Listings of U.S. ProvisionalApplication 60/212,210 filed Jun. 16, 2000 and U.S. Nonprovisionalapplication 09/880,748 filed Jun. 15, 2001 the contents of which areherein incorporated in their entireties by reference.

The entire disclosure (including the specification, sequence listing,and drawings) of each of the following U.S. Provisional andNon-Provisional Patent Applications and International PatentApplications are herein incorporated by reference in their entireties:U.S. Provisional Application Ser. Nos.: 60/543, 296 Filed Feb. 11, 2004;60/580,347 fi9led Jun. 18, 2004; 60/331,469 filed Nov. 16, 2001;60/340,817 filed Dec. 19, 2001; 60/212,210 filed Jun. 16, 2000;60/240,816 filed Oct. 17 2000; 60/276,248 filed Mar. 16, 2001;60/277,379 filed Mar. 21 2001; 60/293,499 filed May 25, 2001; and U.S.Nonprovisional application Ser. Nos. 10/293,418 and 09/880,748 filedJun. 15, 2001; and International Patent Application Serial No.PCT/US01/19110 filed Jun. 15, 2001.

TABLE 1 scFvs that Immunospecifically Bind to B Lymphocyte StimulatorscFv SEQ AAs of AAs of AAs of Clone ID ID NO AAs of VL VL CDR1 VL CDR2VL CDR3 AAs of VH I003F12S 1 138-248 160-173 189-195 228-237 1-122I006D08 2 141-249 163-173 189-195 228-238 1-123 I008A11 3 144-254166-179 195-201 234-243 1-128 I017D10 4 148-255 169-179 195-201 234-2441-132 I022D01 5 142-249 163-173 189-195 228-238 1-126 I031F02 6 138-251160-173 189-195 228-240 1-121 I050A12 7 142-250 164-174 190-196 229-2391-124 I051C04 8 146-256 168-181 197-203 236-245 1-129 I050B11 9 143-251166-177 193-199 232-240 1-125 I050B11-01 10 143-251 166-177 193-199232-240 1-125 I050B11-02 11 143-251 166-177 193-199 232-240 1-125I050B11-03 12 143-251 166-177 193-199 232-240 1-125 I050B11-04 13143-251 166-177 193-199 232-240 1-125 I050B11-05 14 143-251 166-177193-199 232-240 1-125 I050B11-06 15 143-251 166-177 193-199 232-2401-125 I050B11-07 16 143-251 166-177 193-199 232-240 1-125 I050B11-08 17143-251 166-177 193-199 232-240 1-125 I050B11-09 18 143-251 166-177193-199 232-240 1-125 I050B11-10 19 143-251 166-177 193-199 232-2401-125 I050B11-11 20 143-251 166-177 193-199 232-240 1-125 I050B11-12 21143-251 166-177 193-199 232-240 1-125 I050B11-13 22 143-251 166-177193-199 232-240 1-125 I050B11-14 23 143-251 166-177 193-199 232-2401-125 I050B11-15 24 143-251 166-177 193-199 232-240 1-125 I050B11-16 25143-251 166-177 193-199 232-240 1-125 I050B11-17 26 143-251 166-177193-199 232-240 1-125 I050B11-18 27 143-251 166-177 193-199 232-2401-125 I050B11-19 28 143-251 166-177 193-199 232-240 1-125 I050B11-20 29143-251 166-177 193-199 232-240 1-125 I050B11-21 30 143-251 166-177193-199 232-240 1-125 I050B11-22 31 143-251 166-177 193-199 232-2401-125 I050B11-23 32 143-251 166-177 193-199 232-240 1-125 I050B11-24 33143-251 166-177 193-199 232-240 1-125 I050B11-25 34 143-251 166-177193-199 232-240 1-125 I050B11-26 35 143-251 166-177 193-199 232-2401-125 I050B11-27 36 143-251 166-177 193-199 232-240 1-125 I050B11-28 37143-251 166-177 193-199 232-240 1-125 I093D03 38 143-251 166-177 193-199232-240 1-125 I093D09 39 143-251 166-177 193-199 232-240 1-125 I093G0840 143-251 166-177 193-199 232-240 1-125 I097D11 41 143-251 166-177193-199 232-240 1-125 I101A04 42 143-251 166-177 193-199 232-240 1-125I101B01 43 143-251 166-177 193-199 232-240 1-125 I102A02 44 143-251166-177 193-199 232-240 1-125 I102E01 45 143-251 166-177 193-199 232-2401-125 I102G06 46 143-251 166-177 193-199 232-240 1-125 I087A07 47143-251 166-177 193-199 232-240 1-125 I087A08 48 142-250 165-176 192-198231-239 1-124 I087A09 49 142-250 165-176 192-198 231-239 1-124 I087B0250 142-250 165-176 192-198 231-239 1-124 I087B03 51 142-250 165-176192-198 231-239 1-124 I087B04 52 142-250 165-176 192-198 231-239 1-124I087B05 53 142-250 165-176 192-198 231-239 1-124 I087B06 54 142-250165-176 192-198 231-239 1-124 I087B08 55 143-251 166-177 193-199 232-2401-125 I087B09 56 142-250 165-176 192-198 231-239 1-124 I087C02 57142-250 165-176 192-198 231-239 1-124 I087C05 58 142-250 165-176 192-198231-239 1-124 I087C06 59 142-250 165-176 192-198 231-239 1-124 I087C0760 142-250 165-176 192-198 231-239 1-124 I087C08 61 143-251 166-177193-199 232-240 1-125 I087D01 62 142-250 165-176 192-198 231-239 1-124I087B02 63 142-250 165-176 192-198 231-239 1-124 I087D03 64 142-251166-177 193-199 232-240 1-125 I087D05 65 143-251 166-177 193-199 232-2401-125 I087D07 66 142-250 165-176 192-198 231-239 1-124 I087D09 67143-251 166-177 193-199 232-240 1-125 I087E04 68 143-251 166-177 193-199232-240 1-125 I087E05 69 142-250 165-176 192-198 231-239 1-124 I087E1070 143-251 166-177 193-199 232-240 1-125 I087F02 71 143-251 166-177193-199 232-240 1-125 I087F04 72 143-251 166-177 193-199 232-240 1-125I087F05 73 142-250 165-176 192-198 231-239 1-124 I087F07 74 142-250165-176 192-198 231-239 1-124 I087F08 75 142-250 165-176 192-198 231-2391-124 I087F09 76 142-250 165-176 192-198 231-239 1-124 I087G05 77142-250 165-176 192-198 231-239 1-124 I087G06 78 142-250 165-176 192-198231-239 1-124 I087G07 79 143-251 166-177 193-199 232-240 1-125 I087G0980 142-250 165-176 192-198 231-239 1-124 I087G10 81 142-250 165-176192-198 231-239 1-124 I087H02 82 137-244 160-170 186-192 225-233 1-121I088A01 83 143-251 166-177 193-199 232-240 1-125 I088A03 84 143-251166-177 193-199 232-240 1-125 I088A04 85 143-251 166-177 193-199 232-2401-125 I088A08 86 143-251 166-177 193-199 232-240 1-125 I088A09 87143-251 166-177 193-199 232-240 1-125 I088A10 88 143-251 166-177 193-199232-240 1-125 I088A11 89 143-251 166-177 193-199 232-240 1-125 I088A1290 143-251 166-177 193-199 232-240 1-125 I088B01 91 143-251 166-177193-199 232-240 1-125 I088B02 92 143-251 166-177 193-199 232-240 1-125I088B03 93 143-251 166-177 193-199 232-240 1-125 I088B05 94 143-251166-177 193-199 232-240 1-125 I088B06 95 143-251 166-177 193-199 232-2401-125 I088B07 96 143-251 166-177 193-199 232-240 1-125 I088B08 97143-251 166-177 193-199 232-240 1-125 I088B09 98 143-251 166-177 193-199232-240 1-125 I088B10 99 143-251 166-177 193-199 232-240 1-125 I088B12100 142-250 165-176 192-198 231-239 1-124 I088C01 101 143-251 166-177193-199 232-240 1-125 I088C03 102 143-251 166-177 193-199 232-240 1-125I088C09 103 143-251 166-177 193-199 232-240 1-125 I088C12 104 143-251166-177 193-199 232-240 1-125 I088D01 105 143-251 166-177 193-199232-240 1-125 I088D03 106 143-251 166-177 193-199 232-240 1-125 I088D04107 142-250 165-176 192-198 231-239 1-124 I088D07 108 143-251 166-177193-199 232-240 1-125 I088D08 109 143-251 166-177 193-199 232-240 1-125I088D11 110 143-251 166-177 193-199 232-240 1-125 I088E01 111 140-248163-174 190-196 229-237 1-122 I088E02 112 143-251 166-177 193-199232-240 1-125 I088E03 113 143-251 166-177 193-199 232-240 1-125 I088E04114 143-251 166-177 193-199 232-240 1-125 I088E08 115 143-251 166-177193-199 232-240 1-125 I088E10 116 143-251 166-177 193-199 232-240 1-125I088E11 117 143-251 166-177 193-199 232-240 1-125 I088F07 118 143-251166-177 193-199 232-240 1-125 I088G02 119 143-251 166-177 193-199232-240 1-125 I088G03 120 143-251 166-177 193-199 232-240 1-125 I088G07121 143-251 166-177 193-199 232-240 1-125 I088G09 122 143-251 166-177193-199 232-240 1-125 I088G10 123 143-251 166-177 193-199 232-240 1-125I088H05 124 143-251 166-177 193-199 232-240 1-125 I088H07 125 143-251166-177 193-199 232-240 1-125 I092A03 126 143-251 166-177 193-199232-240 1-125 I092A05 127 143-251 166-177 193-199 232-240 1-125 I092A06128 143-251 166-177 193-199 232-240 1-125 I092A08 129 143-251 166-177193-199 232-240 1-125 I092A10 130 143-251 166-177 193-199 232-240 1-125I092A11 131 143-251 166-177 193-199 232-240 1-125 I092B01 132 143-251166-177 193-199 232-240 1-125 I092B02 133 143-251 166-177 193-199232-240 1-125 I092B04 134 143-251 166-177 193-199 232-240 1-125 I092B05135 143-251 166-177 193-199 232-240 1-125 I092B10 136 143-251 166-177193-199 232-240 1-125 I092B12 137 143-251 166-177 193-199 232-240 1-125I092C01 138 143-251 166-177 193-199 232-240 1-125 I092C02 139 143-251166-177 193-199 232-240 1-125 I092C07 140 142-250 165-176 192-198231-239 1-124 I092C08 141 143-251 166-177 193-199 232-240 1-125 I092C12142 143-251 166-177 193-199 232-240 1-125 I092D01 143 143-251 166-177193-199 232-240 1-125 I092D07 144 143-251 166-177 193-199 232-240 1-125I092D09 145 143-251 166-177 193-199 232-240 1-125 I092D10 146 143-251166-177 193-199 232-240 1-125 I092D11 147 143-251 166-177 193-199232-240 1-125 I092E01 148 143-251 166-177 193-199 232-240 1-125 I092E03149 143-251 166-177 193-199 232-240 1-125 I092E04 150 143-251 166-177193-199 232-240 1-125 I092E07 151 143-251 166-177 193-199 232-240 1-125I092E10 152 143-251 166-177 193-199 232-240 1-125 I092E11 153 143-251166-177 193-199 232-240 1-125 I092F01 154 143-251 166-177 193-199232-240 1-125 I092F02 155 143-251 166-177 193-199 232-240 1-125 I092F05156 143-251 166-177 193-199 232-240 1-125 I092F07 157 143-251 166-177193-199 232-240 1-125 I092F08 158 143-251 166-177 193-199 232-240 1-125I092F11 159 143-251 166-177 193-199 232-240 1-125 I092F12 160 143-251166-177 193-199 232-240 1-125 I092G01 161 143-251 166-177 193-199232-240 1-125 I092G05 162 143-251 166-177 193-199 232-240 1-125 I092G10163 143-251 166-177 193-199 232-240 1-125 I092H01 164 137-244 160-170186-192 225-233 1-121 I093A06 165 143-251 166-177 193-199 232-240 1-125I093A09 166 143-251 166-177 193-199 232-240 1-125 I093A11 167 143-251166-177 193-199 232-240 1-125 I093Al2 168 143-251 166-177 193-199232-240 1-125 I093B02 169 143-251 166-177 193-199 232-240 1-125 I093B05170 143-251 166-177 193-199 232-240 1-125 I093B06 171 143-251 166-177193-199 232-240 1-125 I093B09 172 143-251 166-177 193-199 232-240 1-125I093B12 173 143-251 166-177 193-199 232-240 1-125 I093C02 174 143-251166-177 193-199 232-240 1-125 I093C03 175 143-251 166-177 193-199232-240 1-125 I093C05 176 143-251 166-177 193-199 232-240 1-125 I093D05177 143-251 166-177 193-199 232-240 1-125 I093D08 178 143-251 166-177193-199 232-240 1-125 I093D10 179 143-251 166-177 193-199 232-240 1-125I093D12 180 143-251 166-177 193-199 232-240 1-125 I093E01 181 143-251166-177 193-199 232-240 1-125 I093E02 182 143-251 166-177 193-199232-240 1-125 I093E05 183 143-251 166-177 193-199 232-240 1-125 I093E08184 143-251 166-177 193-199 232-240 1-125 I093E10 185 143-251 166-177193-199 232-240 1-125 I093F01 186 143-251 166-177 193-199 232-240 1-125I093F03 187 143-251 166-177 193-199 232-240 1-125 I093F05 188 143-251166-177 193-199 232-240 1-125 I093F08 189 143-251 166-177 193-199232-240 1-125 I093F11 190 143-251 166-177 193-199 232-240 1-125 I093G07191 143-251 166-177 193-199 232-240 1-125 I093G11 192 143-251 166-177193-199 232-240 1-125 I093G12 193 143-251 166-177 193-199 232-240 1-125I093H06 194 143-251 166-177 193-199 232-240 1-125 I094A08 195 143-251166-177 193-199 232-240 1-125 I094B07 196 143-251 166-177 193-199232-240 1-125 I094B08 197 143-251 166-177 193-199 232-240 1-125 I094B12198 143-251 166-177 193-199 232-240 1-125 I094C11 199 143-251 166-177193-199 232-240 1-125 I094C12 200 143-251 166-177 193-199 232-240 1-125I094D06 201 143-251 166-177 193-199 232-240 1-125 I094D07 202 143-251166-177 193-199 232-240 1-125 I094D08 203 143-251 166-177 193-199232-240 1-125 I094D09 204 143-251 166-177 193-199 232-240 1-125 I094D10205 143-251 166-177 193-199 232-240 1-125 I094D11 206 143-251 166-177193-199 232-240 1-125 I094E04 207 143-251 166-177 193-199 232-240 1-125I094E08 208 143-251 166-177 193-199 232-240 1-125 I094F04 209 143-251166-177 193-199 232-240 1-125 I094F05 210 143-251 166-177 193-199232-240 1-125 I094F10 211 143-251 166-177 193-199 232-240 1-125 I094F11212 143-251 166-177 193-199 232-240 1-125 I094F12 213 143-251 166-177193-199 232-240 1-125 I094G06 214 143-251 166-177 193-199 232-240 1-125I094G10 215 143-251 166-177 193-199 232-240 1-125 I095A04 216 143-251166-177 193-199 232-240 1-125 I095Al2 217 143-251 166-177 193-199232-240 1-125 I095B04 218 143-251 166-177 193-199 232-240 1-125 I095B09219 143-251 166-177 193-199 232-240 1-125 I095B10 220 143-251 166-177193-199 232-240 1-125 I095C02 221 143-251 166-177 193-199 232-240 1-125I095C05 222 143-251 166-177 193-199 232-240 1-125 I095C07 223 143-251166-177 193-199 232-240 1-125 I095C08 224 143-251 166-177 193-199232-240 1-125 I095C09 225 143-251 166-177 193-199 232-240 1-125 I095D01226 143-251 166-177 193-199 232-240 1-125 I095D02 227 143-251 166-177193-199 232-240 1-125 I095D03 228 143-251 166-177 193-199 232-240 1-125I095D05 229 143-251 166-177 193-199 232-240 1-125 I095D09 230 143-251166-177 193-199 232-240 1-125 I095E01 231 143-251 166-177 193-199232-240 1-125 I095E05 232 143-251 166-177 193-199 232-240 1-125 I095E12233 143-251 166-177 193-199 232-240 1-125 I095F06 234 143-251 166-177193-199 232-240 1-125 I095F09 235 143-251 166-177 193-199 232-240 1-125I095G06 236 143-251 166-177 193-199 232-240 1-125 I095G09 237 143-251166-177 193-199 232-240 1-125 I095G11 238 143-251 166-177 193-199232-240 1-125 I096A01 239 143-251 166-177 193-199 232-240 1-125 I096A10240 143-251 166-177 193-199 232-240 1-125 I096B01 241 143-251 166-177193-199 232-240 1-125 I096B03 242 143-251 166-177 193-199 232-240 1-125I096C01 243 143-251 166-177 193-199 232-240 1-125 I096C06 244 143-251166-177 193-199 232-240 1-125 I096C09 245 143-251 166-177 193-199232-240 1-125 I096D01 246 143-251 166-177 193-199 232-240 1-125 I096D02247 143-251 166-177 193-199 232-240 1-125 I096D05 248 143-251 166-177193-199 232-240 1-125 I096D06 249 143-251 166-177 193-199 232-240 1-125I096D09 250 143-251 166-177 193-199 232-240 1-125 I096E02 251 143-251166-177 193-199 232-240 1-125 I096E06 252 143-251 166-177 193-199232-240 1-125 I096E11 253 143-251 166-177 193-199 232-240 1-125 I096F02254 143-251 166-177 193-199 232-240 1-125 I096G01 255 143-251 166-177193-199 232-240 1-125 I096G02 256 143-251 166-177 193-199 232-240 1-125I096G05 257 143-251 166-177 193-199 232-240 1-125 I096G07 258 143-251166-177 193-199 232-240 1-125 I096G09 259 143-251 166-177 193-199232-240 1-125 I096G12 260 143-251 166-177 193-199 232-240 1-125 I096H01261 137-244 160-170 186-192 225-233 1-121 I097A04 262 143-251 166-177193-199 232-240 1-125 I097A06 263 143-251 166-177 193-199 232-240 1-125I097A09 264 143-251 166-177 193-199 232-240 1-125 I097B02 265 143-251166-177 193-199 232-240 1-125 I097B09 266 143-251 166-177 193-199232-240 1-125 I097B10 267 143-251 166-177 193-199 232-240 1-125 I097B11268 143-251 166-177 193-199 232-240 1-125 I097C05 269 143-251 166-177193-199 232-240 1-125 I097C09 270 143-251 166-177 193-199 232-240 1-125I097C11 271 143-251 166-177 193-199 232-240 1-125 I097D05 272 143-251166-177 193-199 232-240 1-125 I097D06 273 143-251 166-177 193-199232-240 1-125 I097E01 274 143-251 166-177 193-199 232-240 1-125 I097E04275 143-251 166-177 193-199 232-240 1-125 I097E08 276 143-251 166-177193-199 232-240 1-125 I097E09 277 143-251 166-177 193-199 232-240 1-125I097F09 278 143-251 166-177 193-199 232-240 1-125 I097G10 279 143-251166-177 193-199 232-240 1-125 I097H02 280 137-244 160-170 186-192225-233 1-121 I098A04 281 143-251 166-177 193-199 232-240 1-125 I098A05282 143-251 166-177 193-199 232-240 1-125 I098B08 283 143-251 166-177193-199 232-240 1-125 I098C01 284 143-251 166-177 193-199 232-240 1-125I098C04 285 143-251 166-177 193-199 232-240 1-125 I098F11 286 143-251166-177 193-199 232-240 1-125 I098F12 287 143-251 166-177 193-199232-240 1-125 I098G02 288 143-251 166-177 193-199 232-240 1-125 I098G12289 143-251 166-177 193-199 232-240 1-125 I098H05 290 143-251 166-177193-199 232-240 1-125 I101A01 291 143-251 166-177 193-199 232-240 1-125I101B04 292 143-251 166-177 193-199 232-240 1-125 I101B06 293 143-251166-177 193-199 232-240 1-125 I101D04 294 143-251 166-177 193-199232-240 1-125 I101D07 295 143-251 166-177 193-199 232-240 1-125 I101E09296 143-251 166-177 193-199 232-240 1-125 I101E12 297 143-251 166-177193-199 232-240 1-125 I101G02 298 143-251 166-177 193-199 232-240 1-125I101G11 299 143-251 166-177 193-199 232-240 1-125 I102C03 300 143-251166-177 193-199 232-240 1-125 I102E09 301 143-251 166-177 193-199232-240 1-125 I102F02 302 143-251 166-177 193-199 232-240 1-125 I102G08303 143-251 166-177 193-199 232-240 1-125 I102G09 304 143-251 166-177193-199 232-240 1-125 I106A09 305 143-251 166-177 193-199 232-240 1-125I106B02 306 143-251 166-177 193-199 232-240 1-125 I106B06 307 143-251166-177 193-199 232-240 1-125 I106C07 308 143-251 166-177 193-199232-240 1-125 I106E05 309 143-251 166-177 193-199 232-240 1-125 I106E12310 143-251 166-177 193-199 232-240 1-125 I106G01 311 143-251 166-177193-199 232-240 1-125 I106G03 312 143-251 166-177 193-199 232-240 1-125I109B06 313 143-251 166-177 193-199 232-240 1-125 I109D12 314 143-251166-177 193-199 232-240 1-125 I109E12 315 143-251 166-177 193-199232-240 1-125 I109G06 316 143-251 166-177 193-199 232-240 1-125 I109H04317 143-251 166-177 193-199 232-240 1-125 I110B03 318 143-251 166-177193-199 232-240 1-125 I112D09 319 143-251 166-177 193-199 232-240 1-125I112F10 320 143-251 166-177 193-199 232-240 1-125 I089F12 321 141-249163-173 189-195 228-238 1-123 I105E12 322 141-249 163-173 189-195228-238 1-123 I108D08 323 141-249 163-173 189-195 228-238 1-123 I108E06324 141-249 163-173 189-195 228-238 1-123 I113E07 325 141-249 163-173189-195 228-238 1-123 I114G05 326 141-249 163-173 189-195 228-238 1-123I116A01 327 141-249 163-173 189-195 228-238 1-123 I116A09 328 141-249163-173 189-195 228-238 1-123 I116C11 329 141-249 163-173 189-195228-238 1-123 I085A01 330 141-249 163-173 189-195 228-238 1-123 I085A02331 141-249 163-173 189-195 228-238 1-123 I085A03 332 141-249 163-173189-195 228-238 1-123 I085A04 333 141-249 163-173 189-195 228-238 1-123I085A05 334 141-249 163-173 189-195 228-238 1-123 I085A06 335 141-249163-173 189-195 228-238 1-123 I085A07 336 141-249 163-173 189-195228-238 1-123 I085A09 337 140-248 162-172 188-194 227-237 1-122 I085A10338 141-249 163-173 189-195 228-238 1-123 I085A11 339 140-248 162-172188-194 227-237 1-122 I085B01 340 141-249 163-173 189-195 228-238 1-123I085B02 341 141-249 163-173 189-195 228-238 1-123 I085B03 342 141-249163-173 189-195 228-238 1-123 I085B04 343 141-249 163-173 189-195228-238 1-123 I085B05 344 141-249 163-173 189-195 228-238 1-123 I085B06345 141-249 163-173 189-195 228-238 1-123 I085B07 346 141-249 163-173189-195 228-238 1-123 I085B10 347 140-248 162-172 188-194 227-237 1-122I085B12 348 141-249 163-173 189-195 228-238 1-123 I085C02 349 141-249163-173 189-195 228-238 1-123 I085C03 350 141-249 163-173 189-195228-238 1-123 I085C05 351 141-249 163-173 189-195 228-238 1-123 I085C06352 141-249 163-173 189-195 228-238 1-123 I085C07 353 141-249 163-173189-195 228-238 1-123 I085C09 354 140-248 162-172 188-194 227-237 1-122I085C10 355 141-249 163-173 189-195 228-238 1-123 I085C12 356 141-249163-173 189-195 228-238 1-123 I085D01 357 141-249 163-173 189-195228-238 1-123 I085D02 358 141-249 163-173 189-195 228-238 1-123 I085D03359 141-249 163-173 189-195 228-238 1-123 I085D04 360 141-249 163-173189-195 228-238 1-123 I085D06 361 141-249 163-173 189-195 228-238 1-123I085D07 362 141-249 163-173 189-195 228-238 1-123 I085D08 363 141-249163-173 189-195 228-238 1-123 I085D09 364 141-249 163-173 189-195228-238 1-123 I085D10 365 141-249 163-173 189-195 228-238 1-123 I085D11366 141-249 163-173 189-195 228-238 1-123 I085D12 367 141-249 163-173189-195 228-238 1-123 I085E01 368 141-249 163-173 189-195 228-238 1-123I085E02 369 141-249 163-173 189-195 228-238 1-123 I085E07 370 141-249163-173 189-195 228-238 1-123 I085E08 371 141-249 163-173 189-195228-238 1-123 I085E09 372 141-249 163-173 189-195 228-238 1-123 I085E10373 141-249 163-173 189-195 228-238 1-123 I085E11 374 141-249 163-173189-195 228-238 1-123 I085E12 375 141-249 163-173 189-195 228-238 1-123I085F01 376 141-249 163-173 189-195 228-238 1-123 I085F02 377 141-249163-173 189-195 228-238 1-123 I085F03 378 141-249 163-173 189-195228-238 1-123 I085F04 379 141-249 163-173 189-195 228-238 1-123 I085F05380 141-249 163-173 189-195 228-238 1-123 I085F06 381 141-249 163-173189-195 228-238 1-123 I085F07 382 141-249 163-173 189-195 228-238 1-123I085F09 383 141-249 163-173 189-195 228-238 1-123 I085F10 384 141-249163-173 189-195 228-238 1-123 I085F11 385 141-249 163-173 189-195228-238 1-123 I085F12 386 141-249 163-173 189-195 228-238 1-123 I085G01387 141-249 163-173 189-195 228-238 1-123 I085G02 388 141-249 163-173189-195 228-238 1-123 I085G03 389 141-249 163-173 189-195 228-238 1-123I085G04 390 141-249 163-173 189-195 228-238 1-123 I085G07 391 141-249163-173 189-195 228-238 1-123 I085G08 392 141-249 163-173 189-195228-238 1-123 I085G09 393 141-249 163-173 189-195 228-238 1-123 I085G10394 140-248 162-172 188-194 227-237 1-122 I085G11 395 141-249 163-173189-195 228-238 1-123 I085G12 396 141-249 163-173 189-195 228-238 1-123I085H10 397 142-249 163-173 189-195 228-238 1-126 I086A03 398 141-249163-173 189-195 228-238 1-123 I086A04 399 141-249 163-173 189-195228-238 1-123 I086A05 400 141-249 163-173 189-195 228-238 1-123 I086A07401 141-249 163-173 189-195 228-238 1-123 I086A09 402 141-249 163-173189-195 228-238 1-123 I086A10 403 141-249 163-173 189-195 228-238 1-123I086A11 404 141-249 163-173 189-195 228-238 1-123 I086Al2 405 141-249163-173 189-195 228-238 1-123 I086B02 406 141-249 163-173 189-195228-238 1-123 I086B03 407 141-249 163-173 189-195 228-238 1-123 I086B05408 141-249 163-173 189-195 228-238 1-123 I086B06 409 141-249 163-173189-195 228-238 1-123 I086B07 410 141-249 163-173 189-195 228-238 1-123I086B09 411 141-249 163-173 189-195 228-238 1-123 I086B10 412 141-249163-173 189-195 228-238 1-123 I086B11 413 141-249 163-173 189-195228-238 1-123 I086C03 414 141-249 163-173 189-195 228-238 1-123 I086C05415 141-249 163-173 189-195 228-238 1-123 I086C07 416 141-249 163-173189-195 228-238 1-123 I086C08 417 141-249 163-173 189-195 228-238 1-123I086C09 418 141-249 163-173 189-195 228-238 1-123 I086C10 419 141-249163-173 189-195 228-238 1-123 I086C11 420 141-249 163-173 189-195228-238 1-123 I086C12 421 141-249 163-173 189-195 228-238 1-123 I086D01422 141-249 163-173 189-195 228-238 1-123 I086D04 423 141-249 163-173189-195 228-238 1-123 I086D05 424 141-249 163-173 189-195 228-238 1-123I086D06 425 141-249 163-173 189-195 228-238 1-123 I086D07 426 141-249163-173 189-195 228-238 1-123 I086D08 427 141-249 163-173 189-195228-238 1-123 I086D09 428 141-249 163-173 189-195 228-238 1-123 I086D10429 141-249 163-173 189-195 228-238 1-123 I086D11 430 141-249 163-173189-195 228-238 1-123 I086D12 431 141-249 163-173 189-195 228-238 1-123I086E02 432 141-249 163-173 189-195 228-238 1-123 I086E03 433 141-249163-173 189-195 228-238 1-123 I086E05 434 141-249 163-173 189-195228-238 1-123 I086E07 435 141-249 163-173 189-195 228-238 1-123 I086E08436 141-249 163-173 189-195 228-238 1-123 I086E09 437 141-249 163-173189-195 228-238 1-123 I086E10 438 141-249 163-173 189-195 228-238 1-123I086E12 439 141-249 163-173 189-195 228-238 1-123 I086F02 440 141-249163-173 189-195 228-238 1-123 I086F05 441 141-249 163-173 189-195228-238 1-123 I086F08 442 141-249 163-173 189-195 228-238 1-123 I086F09443 141-249 163-173 189-195 228-238 1-123 I086F11 444 141-249 163-173189-195 228-238 1-123 I086G03 445 141-249 163-173 189-195 228-238 1-123I086G04 446 141-249 163-173 189-195 228-238 1-123 I086G05 447 141-249163-173 189-195 228-238 1-123 I086G06 448 141-249 163-173 189-195228-238 1-123 I086G07 449 141-249 163-173 189-195 228-238 1-123 I086G09450 141-249 163-173 189-195 228-238 1-123 I086G10 451 141-249 163-173189-195 228-238 1-123 I086H05 452 141-249 163-173 189-195 228-238 1-123I089A01 453 140-248 162-172 188-194 227-237 1-122 I089A03 454 141-249163-173 189-195 228-238 1-123 I089A06 455 141-249 163-173 189-195228-238 1-123 I089A07 456 141-249 163-173 189-195 228-238 1-123 I089A08457 141-249 163-173 189-195 228-238 1-123 I089A10 458 141-249 163-173189-195 228-238 1-123 I089A11 459 141-249 163-173 189-195 228-238 1-123I089B01 460 141-249 163-173 189-195 228-238 1-123 I089B02 461 141-249163-173 189-195 228-238 1-123 I089B03 462 141-249 163-173 189-195228-238 1-123 I089B04 463 141-249 163-173 189-195 228-238 1-123 I089B05464 141-249 163-173 189-195 228-238 1-123 I089B06 465 141-249 163-173189-195 228-238 1-123 I089B07 466 141-249 163-173 189-195 228-238 1-123I089B08 467 141-249 163-173 189-195 228-238 1-123 I089B09 468 141-249163-173 189-195 228-238 1-123 I089B10 469 141-249 163-173 189-195228-238 1-123 I089B11 470 141-249 163-173 189-195 228-238 1-123 I089C01471 141-249 163-173 189-195 228-238 1-123 I089C02 472 141-249 163-173189-195 228-238 1-123 I089C03 473 141-249 163-173 189-195 228-238 1-123I089C05 474 141-249 163-173 189-195 228-238 1-123 I089C06 475 141-249163-173 189-195 228-238 1-123 I089C07 476 141-249 163-173 189-195228-238 1-123 I089C09 477 141-249 163-173 189-195 228-238 1-123 I089D01478 141-249 163-173 189-195 228-238 1-123 I089D02 479 141-249 163-173189-195 228-238 1-123 I089D03 480 141-249 163-173 189-195 228-238 1-123I089D04 481 141-249 163-173 189-195 228-238 1-123 I089D05 482 141-249163-173 189-195 228-238 1-123 I089D07 483 141-249 163-173 189-195228-238 1-123 I089D08 484 140-248 162-172 188-194 227-237 1-122 I089D09485 141-249 163-173 189-195 228-238 1-123 I089D11 486 141-249 163-173189-195 228-238 1-123 I089E01 487 141-249 163-173 189-195 228-238 1-123I089E02 488 141-249 163-173 189-195 228-238 1-123 I089E03 489 141-249163-173 189-195 228-238 1-123 I089E04 490 141-249 163-173 189-195228-238 1-123 I089E06 491 141-249 163-173 189-195 228-238 1-123 I089E09492 141-249 163-173 189-195 228-238 1-123 I089E10 493 141-249 163-173189-195 228-238 1-123 I089E11 494 141-249 163-173 189-195 228-238 1-123I089F01 495 141-249 163-173 189-195 228-238 1-123 I089F03 496 141-249163-173 189-195 228-238 1-123 I089F04 497 141-249 163-173 189-195228-238 1-123 I089F05 498 141-249 163-173 189-195 228-238 1-123 I089F06499 141-249 163-173 189-195 228-238 1-123 I089F08 500 141-249 163-173189-195 228-238 1-123 I089F09 501 141-249 163-173 189-195 228-238 1-123I089F10 502 141-249 163-173 189-195 228-238 1-123 I089F11 503 141-249163-173 189-195 228-238 1-123 I089G01 504 141-249 163-173 189-195228-238 1-123 I089G02 505 141-249 163-173 189-195 228-238 1-123 I089G03506 141-249 163-173 189-195 228-238 1-123 I089G05 507 141-249 163-173189-195 228-238 1-123 I089G06 508 141-249 163-173 189-195 228-238 1-123I089G07 509 141-249 163-173 189-195 228-238 1-123 I089G08 510 141-249163-173 189-195 228-238 1-123 I089G11 511 141-249 163-173 189-195228-238 1-123 I089H10 512 142-249 163-173 189-195 228-238 1-126 I090A02513 141-249 163-173 189-195 228-238 1-123 I090A03 514 141-249 163-173189-195 228-238 1-123 I090A04 515 141-249 163-173 189-195 228-238 1-123I090A05 516 141-249 163-173 189-195 228-238 1-123 I090A06 517 141-249163-173 189-195 228-238 1-123 I090A07 518 141-249 163-173 189-195228-238 1-123 I090A08 519 141-249 163-173 189-195 228-238 1-123 I090B01520 141-249 163-173 189-195 228-238 1-123 I090B03 521 141-249 163-173189-195 228-238 1-123 I090B04 522 141-249 163-173 189-195 228-238 1-123I090B05 523 141-249 163-173 189-195 228-238 1-123 I090B06 524 141-249163-173 189-195 228-238 1-123 I090B08 525 141-249 163-173 189-195228-238 1-123 I090B11 526 141-249 163-173 189-195 228-238 1-123 I090B12527 141-249 163-173 189-195 228-238 1-123 I090001 528 141-249 163-173189-195 228-238 1-123 I090002 529 141-249 163-173 189-195 228-238 1-123I090003 530 141-249 163-173 189-195 228-238 1-123 I090005 531 141-249163-173 189-195 228-238 1-123 I090006 532 141-249 163-173 189-195228-238 1-123 I090007 533 141-249 163-173 189-195 228-238 1-123 I090008534 141-249 163-173 189-195 228-238 1-123 I090C10 535 141-249 163-173189-195 228-238 1-123 I090D02 536 141-249 163-173 189-195 228-238 1-123I090D03 537 141-249 163-173 189-195 228-238 1-123 I090D04 538 141-249163-173 189-195 228-238 1-123 I090D05 539 141-249 163-173 189-195228-238 1-123 I090D06 540 141-249 163-173 189-195 228-238 1-123 I090D07541 141-249 163-173 189-195 228-238 1-123 I090D08 542 141-249 163-173189-195 228-238 1-123 I090D09 543 141-249 163-173 189-195 228-238 1-123I090D12 544 141-249 163-173 189-195 228-238 1-123 I090E04 545 141-249163-173 189-195 228-238 1-123 I090E05 546 141-249 163-173 189-195228-238 1-123 I090E06 547 141-249 163-173 189-195 228-238 1-123 I090E07548 141-249 163-173 189-195 228-238 1-123 I090E09 549 141-249 163-173189-195 228-238 1-123 I090E11 550 141-249 163-173 189-195 228-238 1-123I090E12 551 141-249 163-173 189-195 228-238 1-123 I090F01 552 141-249163-173 189-195 228-238 1-123 I090F02 553 141-249 163-173 189-195228-238 1-123 I090F03 554 141-249 163-173 189-195 228-238 1-123 I090F04555 141-249 163-173 189-195 228-238 1-123 I090F05 556 141-249 163-173189-195 228-238 1-123 I090F06 557 141-249 163-173 189-195 228-238 1-123I090F07 558 141-249 163-173 189-195 228-238 1-123 I090F08 559 141-249163-173 189-195 228-238 1-123 I090F09 560 141-249 163-173 189-195228-238 1-123 I090F10 561 141-249 163-173 189-195 228-238 1-123 I090F11562 141-249 163-173 189-195 228-238 1-123 I090G01 563 141-249 163-173189-195 228-238 1-123 I090G02 564 141-249 163-173 189-195 228-238 1-123I090G04 565 141-249 163-173 189-195 228-238 1-123 I090G05 566 141-249163-173 189-195 228-238 1-123 I090G06 567 141-249 163-173 189-195228-238 1-123 I090G07 568 141-249 163-173 189-195 228-238 1-123 I090G08569 141-249 163-173 189-195 228-238 1-123 I090G09 570 141-249 163-173189-195 228-238 1-123 I090G10 571 141-249 163-173 189-195 228-238 1-123I090G12 572 141-249 163-173 189-195 228-238 1-123 I091A02 573 141-249163-173 189-195 228-238 1-123 I091A03 574 141-249 163-173 189-195228-238 1-123 I091A06 575 141-249 163-173 189-195 228-238 1-123 I091A11576 141-249 163-173 189-195 228-238 1-123 I091B01 577 141-249 163-173189-195 228-238 1-123 I091B02 578 141-249 163-173 189-195 228-238 1-123I091B04 579 141-249 163-173 189-195 228-238 1-123 I091B05 580 141-249163-173 189-195 228-238 1-123 I091B07 581 141-249 163-173 189-195228-238 1-123 I091B10 582 141-249 163-173 189-195 228-238 1-123 I091B11583 141-249 163-173 189-195 228-238 1-123 I091B12 584 141-249 163-173189-195 228-238 1-123 I091C02 585 141-249 163-173 189-195 228-238 1-123I091C03 586 141-249 163-173 189-195 228-238 1-123 I091C04 587 141-249163-173 189-195 228-238 1-123 I091C05 588 141-249 163-173 189-195228-238 1-123 I091C06 589 141-249 163-173 189-195 228-238 1-123 I091C09590 141-249 163-173 189-195 228-238 1-123 I091C11 591 141-249 163-173189-195 228-238 1-123 I091C12 592 141-249 163-173 189-195 228-238 1-123I091D01 593 141-249 163-173 189-195 228-238 1-123 I091D02 594 141-249163-173 189-195 228-238 1-123 I091D04 595 141-249 163-173 189-195228-238 1-123 I091D05 596 141-249 163-173 189-195 228-238 1-123 I091D06597 140-248 162-172 188-194 227-237 1-122 I091D07 598 141-249 163-173189-195 228-238 1-123 I091D09 599 141-249 163-173 189-195 228-238 1-123I091E01 600 141-249 163-173 189-195 228-238 1-123 I091E02 601 141-249163-173 189-195 228-238 1-123 I091E03 602 141-249 163-173 189-195228-238 1-123 I091E04 603 141-249 163-173 189-195 228-238 1-123 I091E06604 141-249 163-173 189-195 228-238 1-123 I091E07 605 141-249 163-173189-195 228-238 1-123 I091E08 606 141-249 163-173 189-195 228-238 1-123I091E09 607 141-249 163-173 189-195 228-238 1-123 I091E10 608 141-249163-173 189-195 228-238 1-123 I091F01 609 141-249 163-173 189-195228-238 1-123 I091F03 610 141-249 163-173 189-195 228-238 1-123 I091F05611 141-249 163-173 189-195 228-238 1-123 I091F06 612 141-249 163-173189-195 228-238 1-123 I091F07 613 141-249 163-173 189-195 228-238 1-123I091F08 614 141-249 163-173 189-195 228-238 1-123 I091F09 615 141-249163-173 189-195 228-238 1-123 I091F10 616 141-249 163-173 189-195228-238 1-123 I091F11 617 141-249 163-173 189-195 228-238 1-123 I091F12618 141-249 163-173 189-195 228-238 1-123 I091G01 619 141-249 163-173189-195 228-238 1-123 I091G03 620 141-249 163-173 189-195 228-238 1-123I091G04 621 141-249 163-173 189-195 228-238 1-123 I091G05 622 141-249163-173 189-195 228-238 1-123 I091G06 623 140-248 162-172 188-194227-237 1-122 I091G07 624 141-249 163-173 189-195 228-238 1-123 I091G09625 141-249 163-173 189-195 228-238 1-123 I091G10 626 141-249 163-173189-195 228-238 1-123 I091G11 627 141-249 163-173 189-195 228-238 1-123I091G12 628 141-249 163-173 189-195 228-238 1-123 I104A01 629 141-249163-173 189-195 228-238 1-123 I104A07 630 141-249 163-173 189-195228-238 1-123 I104A08 631 141-249 163-173 189-195 228-238 1-123 I104A09632 141-249 163-173 189-195 228-238 1-123 I104A10 633 141-249 163-173189-195 228-238 1-123 I104A11 634 141-249 163-173 189-195 228-238 1-123I104Al2 635 141-249 163-173 189-195 228-238 1-123 I104B02 636 141-249163-173 189-195 228-238 1-123 I104B04 637 141-249 163-173 189-195228-238 1-123 I104B09 638 141-249 163-173 189-195 228-238 1-123 I104B11639 141-249 163-173 189-195 228-238 1-123 I104C01 640 141-249 163-173189-195 228-238 1-123 I104C04 641 141-249 163-173 189-195 228-238 1-123I104C05 642 141-249 163-173 189-195 228-238 1-123 I104C06 643 140-248162-172 188-194 227-237 1-122 I104C07 644 141-249 163-173 189-195228-238 1-123 I104C09 645 141-249 163-173 189-195 228-238 1-123 I104C11646 141-249 163-173 189-195 228-238 1-123 I104D01 647 141-249 163-173189-195 228-238 1-123 I104D02 648 141-249 163-173 189-195 228-238 1-123I104D03 649 141-249 163-173 189-195 228-238 1-123 I104D04 650 141-249163-173 189-195 228-238 1-123 I104D07 651 141-249 163-173 189-195228-238 1-123 I104D08 652 141-249 163-173 189-195 228-238 1-123 I104D09653 141-249 163-173 189-195 228-238 1-123 I104E01 654 141-249 163-173189-195 228-238 1-123 I104E02 655 141-249 163-173 189-195 228-238 1-123I104E03 656 141-249 163-173 189-195 228-238 1-123 I104E05 657 141-249163-173 189-195 228-238 1-123 I104E11 658 141-249 163-173 189-195228-238 1-123 I104E12 659 141-249 163-173 189-195 228-238 1-123 I104F02660 141-249 163-173 189-195 228-238 1-123 I104F03 661 141-249 163-173189-195 228-238 1-123 I104F04 662 141-249 163-173 189-195 228-238 1-123I104F05 663 141-249 163-173 189-195 228-238 1-123 I104F06 664 141-249163-173 189-195 228-238 1-123 I104F07 665 141-249 163-173 189-195228-238 1-123 I104F10 666 141-249 163-173 189-195 228-238 1-123 I104F11667 141-249 163-173 189-195 228-238 1-123 I104F12 668 141-249 163-173189-195 228-238 1-123 I104G04 669 141-249 163-173 189-195 228-238 1-123I104G05 670 141-249 163-173 189-195 228-238 1-123 I104G09 671 141-249163-173 189-195 228-238 1-123 I104G11 672 141-249 163-173 189-195228-238 1-123 I105A02 673 141-249 163-173 189-195 228-238 1-123 I105A03674 141-249 163-173 189-195 228-238 1-123 I105A04 675 141-249 163-173189-195 228-238 1-123 I105A08 676 141-249 163-173 189-195 228-238 1-123I105A09 677 141-249 163-173 189-195 228-238 1-123 I105A11 678 141-249163-173 189-195 228-238 1-123 I105B04 679 141-249 163-173 189-195228-238 1-123 I105B05 680 141-249 163-173 189-195 228-238 1-123 I105B07681 141-249 163-173 189-195 228-238 1-123 I105B08 682 141-249 163-173189-195 228-238 1-123 I105B10 683 141-249 163-173 189-195 228-238 1-123I105B11 684 141-249 163-173 189-195 228-238 1-123 I105B12 685 141-249163-173 189-195 228-238 1-123 I105C02 686 141-249 163-173 189-195228-238 1-123 I105C03 687 141-249 163-173 189-195 228-238 1-123 I105C05688 141-249 163-173 189-195 228-238 1-123 I105C06 689 141-249 163-173189-195 228-238 1-123 I105C08 690 141-249 163-173 189-195 228-238 1-123I105C12 691 141-249 163-173 189-195 228-238 1-123 I105D04 692 141-249163-173 189-195 228-238 1-123 I105D06 693 141-249 163-173 189-195228-238 1-123 I105D08 694 141-249 163-173 189-195 228-238 1-123 I105D09695 141-249 163-173 189-195 228-238 1-123 I105D10 696 141-249 163-173189-195 228-238 1-123 I105D11 697 141-249 163-173 189-195 228-238 1-123I105E01 698 141-249 163-173 189-195 228-238 1-123 I105E06 699 141-249163-173 189-195 228-238 1-123 I105E11 700 141-249 163-173 189-195228-238 1-123 I105F03 701 141-249 163-173 189-195 228-238 1-123 I105F06702 141-249 163-173 189-195 228-238 1-123 I105F07 703 141-249 163-173189-195 228-238 1-123 I105F09 704 141-249 163-173 189-195 228-238 1-123I105F12 705 141-249 163-173 189-195 228-238 1-123 I105G03 706 141-249163-173 189-195 228-238 1-123 I105G08 707 141-249 163-173 189-195228-238 1-123 I105G09 708 141-249 163-173 189-195 228-238 1-123 I105G10709 141-249 163-173 189-195 228-238 1-123 I105G11 710 141-249 163-173189-195 228-238 1-123 I107A01 711 141-249 163-173 189-195 228-238 1-123I107A03 712 141-249 163-173 189-195 228-238 1-123 I107A06 713 141-249163-173 189-195 228-238 1-123 I107A07 714 141-249 163-173 189-195228-238 1-123 I107A09 715 141-249 163-173 189-195 228-238 1-123 I107Al2716 141-249 163-173 189-195 228-238 1-123 I107B02 717 141-249 163-173189-195 228-238 1-123 I107B04 718 141-249 163-173 189-195 228-238 1-123I107B05 719 141-249 163-173 189-195 228-238 1-123 I107C01 720 139-247161-171 187-193 226-236 1-121 I107C02 721 141-249 163-173 189-195228-238 1-123 I107C04 722 141-249 163-173 189-195 228-238 1-123 I107C06723 141-249 163-173 189-195 228-238 1-123 I107C08 724 141-249 163-173189-195 228-238 1-123 I107C10 725 141-249 163-173 189-195 228-238 1-123I107D01 726 141-249 163-173 189-195 228-238 1-123 I107D04 727 141-249163-173 189-195 228-238 1-123 I107D07 728 141-249 163-173 189-195228-238 1-123 I107D12 729 141-249 163-173 189-195 228-238 1-123 I107E01730 141-249 163-173 189-195 228-238 1-123 I107E05 731 141-249 163-173189-195 228-238 1-123 I107E07 732 141-249 163-173 189-195 228-238 1-123I107E09 733 141-249 163-173 189-195 228-238 1-123 I107F01 734 141-249163-173 189-195 228-238 1-123 I107F05 735 141-249 163-173 189-195228-238 1-123 I107F09 736 141-249 163-173 189-195 228-238 1-123 I107F10737 141-249 163-173 189-195 228-238 1-123 I107G01 738 141-249 163-173189-195 228-238 1-123 I107G05 739 141-249 163-173 189-195 228-238 1-123I107H02 740 141-249 163-173 189-195 228-238 1-123 I107H06 741 141-249163-173 189-195 228-238 1-123 I107H09 742 141-249 163-173 189-195228-238 1-123 I107H10 743 141-249 163-173 189-195 228-238 1-123 I108Al2744 141-249 163-173 189-195 228-238 1-123 I108B03 745 141-249 163-173189-195 228-238 1-123 I108B04 746 141-249 163-173 189-195 228-238 1-123I108C09 747 141-249 163-173 189-195 228-238 1-123 I108C11 748 141-249163-173 189-195 228-238 1-123 I108D10 749 141-249 163-173 189-195228-238 1-123 I108D11 750 141-249 163-173 189-195 228-238 1-123 I108D12751 141-249 163-173 189-195 228-238 1-123 I108E01 752 141-249 163-173189-195 228-238 1-123 I108E03 753 141-249 163-173 189-195 228-238 1-123I108E05 754 141-249 163-173 189-195 228-238 1-123 I108E07 755 141-249163-173 189-195 228-238 1-123 I108E08 756 141-249 163-173 189-195228-238 1-123 I108E09 757 141-249 163-173 189-195 228-238 1-123 I108E10758 141-249 163-173 189-195 228-238 1-123 I108E11 759 141-249 163-173189-195 228-238 1-123 I108F10 760 141-249 163-173 189-195 228-238 1-123I108F12 761 141-249 163-173 189-195 228-238 1-123 I108G01 762 141-249163-173 189-195 228-238 1-123 I108G02 763 141-249 163-173 189-195228-238 1-123 I108G07 764 141-249 163-173 189-195 228-238 1-123 I108G10765 141-249 163-173 189-195 228-238 1-123 I108G11 766 141-249 163-173189-195 228-238 1-123 I108G12 767 141-249 163-173 189-195 228-238 1-123I108H01 768 141-249 163-173 189-195 228-238 1-123 I108H02 769 141-249163-173 189-195 228-238 1-123 I108H06 770 141-249 163-173 189-195228-238 1-123 I108H08 771 141-249 163-173 189-195 228-238 1-123 I111A06772 141-249 163-173 189-195 228-238 1-123 I111B12 773 141-249 163-173189-195 228-238 1-123 I111C01 774 141-249 163-173 189-195 228-238 1-123I111D06 775 141-249 163-173 189-195 228-238 1-123 I111E04 776 141-249163-173 189-195 228-238 1-123 I111E10 777 141-249 163-173 189-195228-238 1-123 I111E11 778 141-250 163-173 189-195 229-239 1-123 I111E12779 141-249 163-173 189-195 228-238 1-123 I111F07 780 141-249 163-173189-195 228-238 1-123 I111G02 781 141-249 163-173 189-195 228-238 1-123I111H10 782 141-249 163-173 189-195 228-238 1-123 I113A04 783 141-249163-173 189-195 228-238 1-123 I113Al2 784 141-249 163-173 189-195228-238 1-123 I113B06 785 141-249 163-173 189-195 228-238 1-123 I113C06786 141-249 163-173 189-195 228-238 1-123 I113G04 787 141-249 163-173189-195 228-238 1-123 I113G05 788 141-249 163-173 189-195 228-238 1-123I113G10 789 141-249 163-173 189-195 228-238 1-123 I113G11 790 141-249163-173 189-195 228-238 1-123 I113H06 791 141-249 163-173 189-195228-238 1-123 I113H07 792 141-249 163-173 189-195 228-238 1-123 I113H09793 141-249 163-173 189-195 228-238 1-123 I114C04 794 141-249 163-173189-195 228-238 1-123 I114C12 795 141-249 163-173 189-195 228-238 1-123I114D04 796 141-249 163-173 189-195 228-238 1-123 I114D06 797 141-249163-173 189-195 228-238 1-123 I114D10 798 141-249 163-173 189-195228-238 1-123 I114E01 799 141-249 163-173 189-195 228-238 1-123 I114E02800 141-249 163-173 189-195 228-238 1-123 I114E03 801 141-249 163-173189-195 228-238 1-123 I114E11 802 141-249 163-173 189-195 228-238 1-123I114H01 803 141-249 163-173 189-195 228-238 1-123 I114H06 804 141-249163-173 189-195 228-238 1-123 I114H09 805 141-249 163-173 189-195228-238 1-123 I115A02 806 141-249 163-173 189-195 228-238 1-123 I115A07807 141-249 163-173 189-195 228-238 1-123 I115B10 808 141-249 163-173189-195 228-238 1-123 I115C05 809 141-249 163-173 189-195 228-238 1-123I115C06 810 141-249 163-173 189-195 228-238 1-123 I115C08 811 141-249163-173 189-195 228-238 1-123 I115C12 812 141-249 163-173 189-195228-238 1-123 I115D07 813 141-249 163-173 189-195 228-238 1-123 I115E09814 141-249 163-173 189-195 228-238 1-123 I115F06 815 141-249 163-173189-195 228-238 1-123 I115F07 816 141-249 163-173 189-195 228-238 1-123I115F12 817 141-249 163-173 189-195 228-238 1-123 I115G04 818 141-249163-173 189-195 228-238 1-123 I115G05 819 141-249 163-173 189-195228-238 1-123 I115G08 820 141-249 163-173 189-195 228-238 1-123 I115H04821 141-249 163-173 189-195 228-238 1-123 I115H07 822 141-249 163-173189-195 228-238 1-123 I115H09 823 141-249 163-173 189-195 228-238 1-123I116A07 824 141-249 163-173 189-195 228-238 1-123 I116B01 825 141-249163-173 189-195 228-238 1-123 I116B12 826 141-249 163-173 189-195228-238 1-123 I116C06 827 141-249 163-173 189-195 228-238 1-123 I116D07828 141-249 163-173 189-195 228-238 1-123 I116E02 829 141-249 163-173189-195 228-238 1-123 I116E04 830 141-249 163-173 189-195 228-238 1-123I116F02 831 141-249 163-173 189-195 228-238 1-123 I116F11 832 141-249163-173 189-195 228-238 1-123 I116G05 833 141-249 163-173 189-195228-238 1-123 I001C09 834 143-250 164-174 190-196 229-239 1-123 I006D07835 141-248 162-172 188-194 227-237 1-125 I007B03 836 143-253 165-178194-200 233-242 1-127 I007F11 837 140-250 162-175 191-197 230-239 1-124I007H08 838 144-254 166-179 195-201 234-243 1-128 I008A09 839 146-256168-181 197-203 236-245 1-130 I008B01 840 141-251 163-176 192-198231-240 1-125 I008C02 841 145-255 167-180 196-202 235-244 1-129 I008C03842 143-250 164-174 190-196 229-239 1-127 I008C12 843 146-256 168-181197-203 236-245 1-130 I012A06 844 147-254 169-179 195-201 234-243 1-129I016E05 845 141-251 163-176 192-198 231-240 1-125 I016F02 846 135-245157-170 186-192 225-234 1-119 I016F04 847 141-251 163-176 192-198231-240 1-125 I016H07 848 141-248 162-172 188-194 227-237 1-125 I018C02849 141-251 163-176 192-198 231-240 1-125 I018C10 850 143-250 164-174190-196 229-239 1-127 I018D07 851 143-250 164-174 190-196 229-239 1-127I018H08 852 141-251 163-176 192-198 231-240 1-125 I018H09 853 141-251163-176 192-198 231-240 1-125 I021B05 854 143-253 165-178 194-200233-242 1-127 I022E02 855 141-251 163-176 192-198 231-240 1-125 I026E03856 143-251 165-175 191-197 230-240 1-125 I027A07 857 145-255 167-179195-201 234-244 1-128 I028A06 858 142-253 164-176 192-198 231-242 1-126I029D07 859 141-250 163-176 192-198 231-239 1-125 I029F11 860 143-253165-177 193-199 232-242 1-127 I031C03 861 138-248 160-172 188-194227-237 1-121 I031C07 862 148-258 170-183 199-205 238-247 1-131 I031F09863 145-255 167-179 195-201 234-244 1-127 I031G08 864 148-259 170-182198-204 237-248 1-131 I031G10 865 148-258 170-182 198-204 237-247 1-131I031G11 866 145-255 167-179 195-201 234-244 1-127 I037E07 867 140-250162-175 191-197 230-239 1-124 I037E12 868 140-250 162-175 191-197230-239 1-124 I050A07 869 146-257 168-181 197-203 236-246 1-129 I061D02870 144-254 166-179 195-201 234-243 1-128 I061E07 871 141-251 163-175191-197 230-240 1-125 I061H01 872 146-256 168-181 197-203 236-245 1-130I001A03 873 144-254 166-179 195-201 234-243 1-128 I001A07 874 141-251163-176 192-198 231-240 1-125 I001A08 875 141-251 163-176 192-198231-240 1-125 I001A10 876 141-251 163-176 192-198 231-240 1-125 I001Al2877 141-248 162-172 188-194 227-237 1-125 I001B02 878 137-247 159-171187-193 226-236 1-121 I001B07 879 141-251 163-176 192-198 231-240 1-125I001C06 880 143-253 165-178 194-200 233-242 1-127 I001C08 881 144-254166-179 195-201 234-243 1-128 I001C12 882 141-251 163-176 192-198231-240 1-125 I001D08 883 140-250 162-175 191-197 230-239 1-124 I001D12884 141-251 163-176 192-198 231-240 1-125 I001E05 885 143-253 165-178194-200 233-242 1-127 I001E07 886 141-251 163-176 192-198 231-240 1-125I001G09 887 141-251 163-176 192-198 231-240 1-125 I001H05 888 144-254166-179 195-201 234-243 1-128 I001H08 889 141-251 163-176 192-198231-240 1-125 I003A01 890 141-251 163-176 192-198 231-240 1-124 I003A06891 141-251 163-176 192-198 231-240 1-124 I003A07 892 142-249 163-173189-195 228-238 1-126 I003A10 893 141-248 162-172 188-194 227-237 1-125I003B03 894 141-251 163-176 192-198 231-240 1-124 I003B04 895 140-248162-172 188-194 227-237 1-122 I003B09 896 142-249 163-173 189-195228-238 1-126 I003C01 897 142-252 164-176 192-198 231-241 1-124 I003C02898 142-252 164-176 192-198 231-241 1-125 I003C03 899 142-250 164-174190-196 229-239 1-125 I003C12 900 141-248 162-172 188-194 227-237 1-125I003D04 901 140-250 162-174 190-196 229-239 1-123 I003E05 902 142-253164-176 192-198 231-242 1-125 I003F01 903 141-251 163-176 192-198231-240 1-124 I003F02 904 140-251 162-175 191-197 230-240 1-122 I003G01905 145-254 168-179 195-201 234-243 1-127 I003G05 906 144-255 166-179195-201 234-244 1-127 I003G06 907 146-256 168-181 197-203 236-245 1-129I003G11 908 144-251 165-175 191-197 230-240 1-128 I003H02 909 142-253164-176 192-198 233-242 1-124 I003H05 910 141-251 163-176 192-198231-240 1-124 I003H08 911 142-249 163-173 189-195 228-238 1-126 I005A01912 141-249 162-172 188-194 227-238 1-125 I005A02 913 141-248 162-172188-194 227-237 1-125 I005B01 914 141-248 162-172 188-194 227-237 1-125I005B09 915 137-247 159-172 188-194 227-236 1-121 I005C01 916 141-248162-172 188-194 227-237 1-125 I005D02 917 141-251 163-176 192-198231-240 1-125 I005D03 918 142-249 165-175 191-197 230-238 1-126 I005E01919 142-249 165-175 191-197 230-238 1-126 I005E08 920 141-248 162-172188-194 227-237 1-125 I005F01 921 142-248 164-174 190-196 229-238 1-124I005F02 922 144-251 167-177 193-199 232-240 1-128 I005F04 923 137-247159-172 188-194 227-236 1-121 I005F08 924 140-247 161-171 187-193226-236 1-124 I005G01 925 141-251 163-176 192-198 231-240 1-125 I005G08926 142-249 165-175 191-197 230-238 1-126 I005H02 927 140-247 161-171187-193 226-236 1-124 I006B01 928 139-246 160-170 186-192 225-235 1-123I006C09 929 143-253 165-177 193-199 232-242 1-127 I006D09 930 141-251163-176 192-198 231-240 1-125 I006E01 931 143-253 165-178 194-200233-242 1-127 I006E07 932 143-250 166-176 192-198 231-239 1-127 I006F01933 140-250 162-175 191-197 230-239 1-124 I006F02 934 142-253 164-176192-198 231-242 1-126 I006F07 935 143-253 165-177 193-199 232-242 1-127I006G01 936 146-253 169-179 195-201 234-242 1-130 I006G04 937 132-239153-163 179-185 218-228 1-116 I006H01 938 146-253 167-177 193-199232-242 1-130 I006H02 939 143-253 165-177 193-199 232-242 1-127 I007A01940 141-251 163-176 192-198 231-240 1-125 I007A08 941 139-249 161-174190-196 229-238 1-123 I007A11 942 140-250 162-175 191-197 230-239 1-124I007Al2 943 144-251 165-175 191-197 230-240 1-128 I007B04 944 141-251163-176 192-198 231-240 1-125 I007C04 945 141-251 163-176 192-198231-240 1-125 I007C08 946 142-249 163-173 189-195 228-238 1-126 I007C12947 140-250 162-175 191-197 230-239 1-124 I007D07 948 140-247 161-171187-193 226-236 1-124 I007D08 949 144-251 165-175 191-197 230-240 1-128I007E03 950 141-248 162-172 188-194 227-237 1-125 I007E10 951 141-251163-176 192-198 231-240 1-125 I007E11 952 144-251 165-175 191-197230-240 1-128 I007F06 953 141-248 162-172 188-194 227-237 1-125 I007F08954 143-253 165-178 194-200 233-242 1-127 I007G07 955 141-251 163-176192-198 231-240 1-125 I007G09 956 142-252 164-177 193-199 232-241 1-126I007G10 957 142-249 163-173 189-195 228-238 1-126 I007H07 958 147-257169-182 198-204 237-246 1-131 I007H11 959 141-248 162-172 188-194227-237 1-125 I008A02 960 141-251 163-176 192-198 231-240 1-125 I008A05961 141-251 163-176 192-198 231-240 1-125 I008A06 962 141-251 163-176192-198 231-240 1-125 I008A07 963 142-249 163-173 189-195 228-238 1-126I008Al2 964 140-250 162-175 191-197 230-239 1-124 I008B02 965 141-248162-172 188-194 227-237 1-125 I008B04 966 143-253 165-178 194-200233-242 1-127 I008B05 967 141-248 162-172 188-194 227-237 1-125 I008B06968 141-251 163-176 192-198 231-240 1-125 I008B07 969 140-247 163-173189-195 228-236 1-124 I008B10 970 141-248 162-172 188-194 227-237 1-125I008B11 971 144-254 166-179 195-201 234-243 1-128 I008C06 972 141-251163-176 192-198 231-240 1-125 I008C08 973 149-259 171-183 199-205238-248 1-133 I008C09 974 142-249 163-173 189-195 228-238 1-126 I008D01975 141-251 163-176 192-198 231-240 1-125 I008D02 976 141-251 163-176192-198 231-240 1-125 I008D03 977 144-254 166-179 195-201 234-243 1-128I008D04 978 141-251 163-176 192-198 231-240 1-125 I008D05 979 141-251163-176 192-198 231-240 1-125 I008D06 980 141-248 162-172 188-194227-237 1-125 I008D07 981 144-254 166-179 195-201 234-243 1-128 I008D08982 144-251 165-175 191-197 230-240 1-128 I008D12 983 144-254 166-179195-201 234-243 1-128 I008E01 984 141-248 162-172 188-194 227-237 1-125I008E02 985 137-247 159-172 188-194 227-236 1-121 I008E03 986 141-251163-176 192-198 231-240 1-125 I008E04 987 141-248 162-172 188-194227-237 1-125 I008E08 988 141-252 163-175 191-197 230-241 1-125 I008E09989 143-253 165-178 194-200 233-242 1-127 I008E12 990 141-251 163-176192-198 231-240 1-125 I008F03 991 141-251 163-176 192-198 231-240 1-125I008F06 992 141-251 163-176 192-198 231-240 1-125 I008F07 993 143-250164-174 190-196 229-239 1-127 I008F08 994 143-253 165-178 194-200233-242 1-127 I008F09 995 133-243 155-168 184-190 223-232 1-127 I008F10996 140-247 161-171 187-193 226-236 1-124 I008F11 997 144-251 165-175191-197 230-240 1-128 I008G02 998 141-251 163-176 192-198 231-240 1-125I008G03 999 140-247 161-171 187-193 226-236 1-124 I008G04 1000 143-253165-178 194-200 233-242 1-127 I008G05 1001 144-254 166-179 195-201234-243 1-128 I008G11 1002 136-246 158-171 187-193 226-235 1-120 I008G121003 143-253 165-178 194-200 233-242 1-127 I008H02 1004 141-248 164-174190-196 229-237 1-125 I008H03 1005 141-251 163-176 192-198 231-240 1-125I008H06 1006 143-253 165-178 194-200 233-242 1-127 I008H09 1007 143-253165-178 194-200 233-242 1-127 I008H11 1008 141-248 164-174 190-196229-237 1-125 I012B03 1009 141-249 163-175 191-197 230-238 1-124 I012B061010 141-251 163-176 192-198 231-240 1-124 I012B10 1011 141-251 163-175191-197 230-240 1-124 I012C03 1012 143-255 165-178 194-200 233-244 1-126I012C06 1013 141-251 163-176 192-198 231-240 1-124 I012C09 1014 142-250164-174 190-196 229-239 1-124 I012D12 1015 146-256 168-180 196-202235-245 1-129 I012E07 1016 142-252 164-176 192-198 231-241 1-124 I012E081017 140-250 162-174 190-196 229-239 1-123 I012E09 1018 141-247 163-173189-195 228-236 1-124 I012F05 1019 141-249 163-173 189-195 228-238 1-124I012F12 1020 142-251 164-176 192-198 231-240 1-124 I012G03 1021 142-252164-176 192-198 231-241 1-124 I012G05 1022 141-250 163-173 189-195228-239 1-123 I012G10 1023 140-251 162-175 191-197 230-240 1-123 I012H091024 141-249 163-173 189-195 228-238 1-124 I013A10 1025 148-259 170-182198-204 237-248 1-131 I013Al2 1026 149-256 171-181 197-203 236-245 1-131I013B04 1027 149-256 172-182 198-204 237-245 1-131 I013B09 1028 149-257171-181 197-203 236-246 1-131 I013C02 1029 148-258 170-182 198-204237-247 1-131 I013C04 1030 139-249 161-173 189-195 228-238 1-121 I013D021031 138-248 160-173 189-195 228-237 1-121 I013D03 1032 148-259 170-183199-205 238-248 1-131 I013D10 1033 146-257 168-181 197-203 236-246 1-129I013E02 1034 148-259 170-183 199-205 238-248 1-131 I013E05 1035 139-249162-173 189-195 228-238 1-121 I013E09 1036 148-260 170-183 199-205238-249 1-131 I013F03 1037 138-248 160-172 188-194 227-237 1-121 I013F041038 148-258 170-182 198-204 237-247 1-131 I013F07 1039 147-260 170-185201-207 240-249 1-129 I013F09 1040 138-248 160-172 188-194 227-237 1-121I013F10 1041 148-259 170-183 199-205 238-248 1-131 I013H04 1042 148-258170-182 198-204 237-247 1-131 I013H07 1043 148-259 170-183 199-205238-248 1-131 I014Al2 1044 143-253 165-178 194-200 233-242 1-127 I014C061045 142-254 164-177 193-200 233-243 1-125 I014C10 1046 141-251 163-176192-198 231-240 1-125 I014C12 1047 141-251 163-176 192-198 231-240 1-125I014E06 1048 142-252 164-176 192-198 231-241 1-124 I014F02 1049 143-251166-176 192-198 231-240 1-125 I016A08 1050 144-251 165-175 191-197230-240 1-128 I016A09 1051 141-251 163-176 192-198 231-240 1-125 I016C021052 141-251 163-176 192-198 231-240 1-125 I016C03 1053 141-251 163-176192-198 231-240 1-125 I016C05 1054 148-255 169-179 195-201 234-244 1-132I016C09 1055 141-251 163-176 192-198 231-240 1-125 I016C11 1056 148-255169-179 195-201 234-244 1-132 I016D10 1057 141-251 163-176 192-198231-240 1-125 I016D11 1058 141-251 163-176 192-198 231-240 1-125 I016E031059 141-251 163-176 192-198 231-240 1-125 I016E04 1060 141-251 163-176192-198 231-240 1-125 I016F03 1061 141-251 163-176 192-198 231-240 1-125I016F11 1062 141-251 163-176 192-198 231-240 1-125 I016G01 1063 141-251163-176 192-198 231-240 1-125 I016G06 1064 141-251 163-176 192-198231-240 1-125 I016G12 1065 148-255 169-179 195-201 234-244 1-132 I016H101066 141-251 163-176 192-198 231-240 1-125 I017A06 1067 141-251 163-176192-198 231-240 1-125 I017A07 1068 141-251 163-176 192-198 231-240 1-125I017A11 1069 140-253 162-175 191-197 233-242 1-124 I017E12 1070 141-251163-176 192-198 231-240 1-125 I017G03 1071 141-251 163-176 192-198231-240 1-125 I017G07 1072 141-251 163-176 192-198 231-240 1-125 I017H011073 141-251 163-176 192-198 231-240 1-125 I018A02 1074 141-251 163-176192-198 231-240 1-125 I018A04 1075 144-254 166-179 195-201 234-243 1-128I018A05 1076 141-251 163-176 192-198 231-240 1-125 I018A11 1077 141-251163-176 192-198 231-240 1-125 I018B02 1078 141-251 163-176 192-198231-240 1-125 I018B08 1079 141-251 163-176 192-198 231-240 1-125 I018C041080 141-251 163-176 192-198 231-240 1-125 I018D02 1081 141-251 163-176192-198 231-240 1-125 I018E06 1082 141-251 163-176 192-198 231-240 1-125I018E08 1083 141-251 163-176 192-198 231-240 1-125 I018F04 1084 141-251163-176 192-198 231-240 1-125 I018G06 1085 141-251 163-176 192-198231-240 1-125 I018H07 1086 141-251 163-176 192-198 231-240 1-125 I019E051087 144-254 166-179 195-201 234-243 1-128 I019F06 1088 144-254 166-179195-201 234-243 1-128 I019G12 1089 143-253 165-178 194-200 233-242 1-127I020D01 1090 137-247 159-171 187-193 226-236 1-121 I020D05 1091 143-253165-178 194-200 233-242 1-127 I020E10 1092 143-253 165-178 194-200233-242 1-127 I020G12 1093 143-253 165-178 194-200 233-242 1-127 I020H061094 143-253 165-178 194-200 233-242 1-127 I020H10 1095 143-253 165-178194-200 233-242 1-127 I021A11 1096 143-253 165-178 194-200 233-242 1-127I021B01 1097 143-253 165-178 194-200 233-242 1-127 I021C11 1098 143-253165-178 194-200 233-242 1-127 I021D12 1099 137-247 159-171 187-193226-236 1-121 I021E10 1100 143-253 165-178 194-200 233-242 1-127 I021G021101 143-253 165-178 194-200 233-242 1-127 I022A08 1102 142-249 163-173189-195 228-238 1-126 I022B01 1103 141-251 163-176 192-198 231-240 1-125I022B10 1104 141-248 164-174 190-196 229-237 1-125 I022C02 1105 142-249163-173 189-195 228-238 1-126 I022C04 1106 141-251 163-176 192-198231-240 1-125 I022C08 1107 141-251 163-176 192-198 231-240 1-125 I022D061108 142-249 163-173 189-195 228-238 1-126 I022E08 1109 142-249 163-173189-195 228-238 1-126 I022F01 1110 142-249 163-173 189-195 228-238 1-126I022F04 1111 142-249 163-173 189-195 228-238 1-126 I022F12 1112 140-247161-171 187-193 226-236 1-124 I022G11 1113 142-249 163-173 189-195228-238 1-126 I023D01 1114 141-251 163-176 192-198 231-240 1-125 I023D041115 142-249 163-173 189-195 228-238 1-126 I024B04 1116 140-247 161-171187-193 226-236 1-124 I024D01 1117 142-249 163-173 189-195 228-238 1-126I024F06 1118 142-249 163-173 189-195 228-238 1-126 I024H01 1119 142-249163-173 189-195 228-238 1-126 I024H07 1120 142-249 163-173 189-195228-238 1-126 I025A01 1121 141-251 163-176 192-198 231-240 1-124 I025A041122 141-251 163-175 191-197 230-240 1-124 I025A07 1123 141-249 163-173189-195 228-238 1-124 I025B01 1124 134-244 156-168 184-190 223-233 1-117I025B10 1125 142-253 164-176 192-198 233-242 1-124 I025B12 1126 141-251163-176 192-198 231-240 1-124 I025C07 1127 141-251 163-176 192-198231-240 1-124 I025D11 1128 142-252 164-176 192-198 231-241 1-124 I025E041129 142-252 164-176 192-198 231-241 1-126 I025E05 1130 141-251 163-175191-197 230-240 1-124 I025E07 1131 142-252 164-176 192-198 231-241 1-124I025E10 1132 141-251 163-176 192-198 231-240 1-125 I025F01 1133 140-251162-175 191-197 230-240 1-123 I025F08 1134 138-248 160-172 188-194227-237 1-121 I025G03 1135 142-252 164-176 192-198 231-241 1-124 I025G081136 141-254 163-176 192-198 231-243 1-124 I025H02 1137 145-255 167-179195-201 234-244 1-128 I026A01 1138 143-249 165-175 191-197 230-238 1-125I026B01 1139 144-254 166-178 194-200 233-243 1-127 I026B06 1140 141-251163-176 192-198 231-240 1-124 I026C06 1141 141-251 163-176 192-198231-240 1-124 I026C10 1142 139-249 161-174 190-196 229-238 1-122 I026C111143 141-251 163-176 192-198 231-240 1-125 I026D09 1144 140-252 162-175191-197 230-241 1-123 I026E04 1145 142-252 164-176 192-198 231-241 1-124I026E06 1146 141-251 163-175 191-197 230-240 1-124 I026E09 1147 141-251163-176 192-198 231-240 1-124 I026F01 1148 141-251 163-176 192-198231-240 1-124 I026F09 1149 141-251 163-176 192-198 231-240 1-124 I026F121150 141-256 163-176 192-202 237-245 1-124 I026G08 1151 141-251 163-176192-198 231-240 1-124 I026G10 1152 141-251 163-176 192-198 231-240 1-124I026G11 1153 144-255 166-179 195-201 234-244 1-127 I026H02 1154 140-251162-175 191-197 230-240 1-123 I026H06 1155 141-251 163-175 191-197230-240 1-124 I026H10 1156 145-255 167-179 195-201 234-244 1-128 I027A091157 141-251 163-176 192-198 231-240 1-124 I027B02 1158 140-250 162-174190-196 229-239 1-123 I027B05 1159 141-250 163-176 192-198 230-239 1-124I027C08 1160 139-249 161-174 190-196 229-238 1-122 I027D02 1161 142-250164-174 190-196 229-239 1-125 I027E03 1162 141-251 163-176 192-198231-240 1-124 I027E05 1163 142-252 164-176 192-198 231-241 1-124 I027F041164 145-252 167-176 192-198 231-241 1-128 I027F05 1165 141-254 163-176192-198 231-243 1-124 I027F11 1166 141-251 163-176 192-198 231-240 1-124I027G06 1167 142-253 164-176 192-198 233-242 1-124 I027G07 1168 142-250164-174 190-196 229-239 1-124 I027H03 1169 142-252 164-176 192-198231-241 1-125 I028A04 1170 143-250 164-174 190-196 229-239 1-127 I028A071171 141-251 163-176 192-198 231-240 1-125 I028B08 1172 141-251 163-176192-198 231-240 1-125 I028B10 1173 148-258 170-183 199-205 238-247 1-132I028C01 1174 142-250 165-175 191-197 230-239 1-126 I028C04 1175 143-253165-178 194-200 233-242 1-127 I028C08 1176 141-251 163-176 192-198231-240 1-125 I028D04 1177 140-247 163-173 189-195 228-236 1-124 I028D051178 141-248 162-172 188-194 227-237 1-125 I028D12 1179 143-250 164-174190-196 229-239 1-127 I028E06 1180 143-253 165-178 194-200 233-242 1-127I028E07 1181 141-248 162-172 188-194 227-237 1-125 I028E08 1182 141-248162-172 188-194 227-237 1-125 I028F06 1183 146-256 168-180 196-202235-245 1-130 I028F08 1184 134-244 156-169 185-191 224-233 1-118 I028G081185 141-251 163-176 192-198 231-240 1-125 I028G09 1186 144-254 166-179195-201 234-243 1-128 I028G10 1187 141-251 163-176 192-198 231-240 1-125I028H02 1188 142-249 165-175 191-197 230-238 1-126 I028H03 1189 148-256169-179 195-201 234-245 1-132 I028H06 1190 145-255 167-180 196-202235-244 1-129 I028H09 1191 140-250 162-175 191-197 230-239 1-124 I029A101192 139-246 160-170 186-192 225-235 1-123 I029Al2 1193 137-247 159-171187-193 226-236 1-121 I029B11 1194 141-251 163-176 192-198 231-240 1-125I029C08 1195 144-254 166-179 195-201 234-243 1-128 I029E10 1196 144-254166-179 195-201 234-243 1-128 I029F08 1197 144-254 166-179 195-201234-243 1-128 I029G08 1198 141-248 162-172 188-194 227-237 1-125 I030A021199 143-253 165-177 193-199 232-242 1-126 I030A03 1200 141-253 163-175191-197 230-242 1-124 I030A04 1201 141-252 163-176 192-198 231-241 1-124I030A05 1202 141-249 163-175 191-197 230-238 1-124 I030A09 1203 140-250162-174 190-196 229-239 1-123 I030Al2 1204 141-249 163-173 189-195228-238 1-123 I030B06 1205 141-249 163-173 189-195 228-238 1-123 I030B081206 141-247 163-173 189-195 228-236 1-124 I030B10 1207 143-251 165-175191-197 230-240 1-125 I030003 1208 140-252 162-175 191-197 230-241 1-123I030006 1209 147-256 169-182 198-204 237-245 1-130 I030008 1210 134-244156-168 184-190 223-233 1-117 I030009 1211 141-251 163-175 191-197230-240 1-124 I030C10 1212 141-250 163-175 191-197 230-239 1-124 I030C111213 140-251 162-175 191-197 230-240 1-123 I030C12 1214 134-244 156-168184-190 223-233 1-117 I030D07 1215 141-249 163-173 189-195 228-238 1-123I030D12 1216 141-251 163-175 191-197 230-240 1-124 I030E02 1217 140-251162-175 191-197 230-240 1-123 I030E05 1218 142-252 164-176 192-198231-241 1-124 I030E07 1219 142-251 165-176 192-198 231-240 1-124 I030E081220 141-251 163-175 191-197 230-240 1-124 I030E09 1221 141-252 163-176192-198 231-241 1-124 I030E10 1222 140-250 162-174 190-196 229-239 1-123I030F02 1223 142-252 164-176 192-198 231-241 1-125 I030F05 1224 141-251163-175 191-197 230-240 1-124 I030F06 1225 140-251 162-175 191-197230-240 1-123 I030F08 1226 141-254 163-176 192-198 231-243 1-124 I030F091227 142-253 164-176 192-198 231-242 1-124 I030F11 1228 140-250 162-174190-196 229-239 1-123 I030F12 1229 141-251 163-175 191-197 230-240 1-124I030G03 1230 141-256 163-176 192-202 237-245 1-124 I030G07 1231 140-251162-175 191-197 230-240 1-123 I030G09 1232 142-251 164-174 190-196229-240 1-124 I030H05 1233 146-255 168-181 197-203 236-244 1-129 I030H061234 148-258 170-182 198-204 239-247 1-130 I030H10 1235 141-253 163-176192-198 231-242 1-124 I030H11 1236 142-252 164-176 192-198 231-241 1-124I031A01 1237 138-248 160-173 189-195 228-237 1-121 I031A03 1238 143-251166-176 192-198 231-240 1-125 I031A08 1239 148-258 170-182 198-204237-247 1-131 I031Al2 1240 147-257 169-181 197-203 236-246 1-130 I031B031241 137-246 159-172 188-194 227-235 1-120 I031B06 1242 143-253 165-177193-199 232-242 1-126 I031B07 1243 148-258 170-182 198-204 237-247 1-131I031B08 1244 149-260 171-183 199-205 238-249 1-131 I031B09 1245 148-258170-182 198-204 237-247 1-131 I031B11 1246 138-248 160-172 188-194227-237 1-121 I031B12 1247 148-259 170-183 199-205 238-248 1-131 I031C011248 138-248 160-172 188-194 227-237 1-121 I031C02 1249 142-253 164-177193-199 232-242 1-125 I031C04 1250 149-260 171-183 199-205 238-249 1-131I031C08 1251 139-248 161-171 187-193 226-237 1-121 I031C11 1252 149-257171-181 197-203 236-246 1-131 I031D01 1253 146-256 168-180 196-202235-245 1-129 I031D04 1254 138-248 160-172 188-194 227-237 1-121 I031D061255 148-258 170-182 198-204 237-247 1-131 I031D08 1256 145-257 167-180196-202 235-246 1-128 I031D09 1257 139-247 161-171 187-193 226-236 1-121I031D11 1258 149-256 171-181 197-203 236-245 1-131 I031D12 1259 146-254168-178 194-200 233-243 1-128 I031E01 1260 148-258 170-182 198-204237-247 1-131 I031E05 1261 149-257 171-181 197-203 236-246 1-131 I031E071262 148-259 170-182 198-204 237-248 1-131 I031E08 1263 148-259 170-183199-205 238-248 1-131 I031E09 1264 139-246 162-173 189-195 228-235 1-121I031E10 1265 148-258 170-182 198-204 237-247 1-131 I031E11 1266 148-258170-182 198-204 237-247 1-131 I031F01 1267 138-248 160-172 188-194227-237 1-121 I031F04 1268 139-246 162-172 188-194 227-235 1-121 I031F061269 137-247 159-171 187-193 226-236 1-119 I031F10 1270 148-259 170-183199-205 238-248 1-131 I031F11 1271 145-255 167-179 195-201 234-244 1-128I031F12 1272 138-249 160-172 188-194 227-238 1-121 I031G01 1273 138-248160-172 188-194 227-237 1-121 I031G03 1274 148-258 170-182 198-204237-247 1-131 I031G05 1275 148-259 170-183 199-205 238-248 1-131 I031G061276 148-258 170-182 198-204 237-247 1-131 I031G07 1277 149-259 171-183199-205 238-248 1-131 I031G09 1278 148-263 170-183 199-209 244-252 1-131I031G12 1279 146-256 168-180 196-202 235-245 1-129 I031H01 1280 138-250160-173 189-195 228-239 1-121 I031H02 1281 143-255 165-178 194-200233-244 1-126 I031H03 1282 148-260 170-183 199-205 238-249 1-131 I031H061283 145-257 167-179 195-201 234-246 1-128 I031H09 1284 145-255 167-179195-201 234-244 1-128 I031H10 1285 144-256 166-179 195-201 234-245 1-127I031H11 1286 136-246 158-170 186-192 225-235 1-119 I033A08 1287 144-254166-179 195-201 234-243 1-128 I033B11 1288 144-254 166-179 195-201234-243 1-128 I033C01 1289 144-254 166-179 195-201 234-243 1-128 I033C081290 142-249 163-173 189-195 228-238 1-126 I033D02 1291 138-245 161-171187-193 226-234 1-122 I033D03 1292 141-251 163-176 192-198 231-240 1-125I033D05 1293 141-248 162-172 188-194 227-237 1-125 I033D11 1294 140-247161-171 187-193 226-236 1-124 I033D12 1295 144-254 166-179 195-201234-243 1-128 I033E01 1296 139-249 161-173 189-195 228-238 1-123 I033E061297 141-251 163-176 192-198 231-240 1-125 I033E11 1298 143-253 165-177193-199 232-242 1-127 I033E12 1299 142-249 163-173 189-195 228-238 1-126I033F03 1300 139-246 160-170 186-192 225-235 1-123 I033F08 1301 145-256167-179 195-201 234-245 1-129 I033F10 1302 144-254 166-179 195-201234-243 1-128 I033F12 1303 134-241 155-165 181-187 220-230 1-118 I033G011304 143-253 165-178 194-200 233-242 1-127 I033G03 1305 142-249 163-173189-195 228-238 1-126 I033G08 1306 141-248 162-172 188-194 227-237 1-125I033H04 1307 140-247 161-171 187-193 226-236 1-124 I037A05 1308 139-246160-170 186-192 225-235 1-123 I037B03 1309 141-251 163-175 191-197230-240 1-125 I037B04 1310 144-251 167-177 193-199 232-240 1-128 I037C041311 142-252 164-177 193-199 232-241 1-126 I037C06 1312 141-249 163-173189-195 228-238 1-125 I037C08 1313 140-250 162-175 191-197 230-239 1-124I037D11 1314 136-246 158-171 187-193 226-235 1-120 I037E06 1315 144-251165-175 191-197 230-240 1-128 I037F04 1316 144-251 165-175 191-197230-240 1-128 I037G01 1317 141-251 163-176 192-198 231-240 1-125 I037G031318 146-256 168-181 197-203 236-245 1-130 I037G10 1319 140-250 162-175191-197 230-239 1-124 I042A07 1320 144-251 167-177 193-199 232-240 1-128I042A10 1321 142-249 165-175 191-197 230-238 1-126 I042B03 1322 140-247161-171 187-193 226-236 1-124 I042B12 1323 141-248 162-172 188-194227-237 1-125 I042D01 1324 136-246 158-171 187-193 226-235 1-120 I042D031325 140-250 162-175 191-197 230-239 1-124 I042D10 1326 142-252 164-177193-199 232-241 1-126 I042E10 1327 147-257 169-182 198-204 237-246 1-131I042E11 1328 140-247 161-171 187-193 226-236 1-124 I042F08 1329 142-252164-177 193-199 232-241 1-126 I042F12 1330 140-247 161-171 187-193226-236 1-124 I042G08 1331 141-248 162-172 188-194 227-237 1-125 I042G101332 141-251 163-176 192-198 231-240 1-125 I042H03 1333 143-253 165-178194-200 233-242 1-127 I043A03 1334 144-254 166-179 195-201 234-243 1-128I043B02 1335 142-249 163-173 189-195 228-238 1-126 I043B03 1336 141-251163-176 192-198 231-240 1-125 I043B06 1337 143-253 165-178 194-200233-242 1-127 I043B07 1338 141-251 163-176 192-198 231-240 1-125 I043B091339 143-253 165-178 194-200 233-242 1-127 I043D11 1340 144-254 166-179195-201 234-243 1-128 I043E05 1341 143-250 164-174 190-196 229-239 1-127I043F01 1342 141-251 163-176 192-198 231-240 1-125 I043F04 1343 141-251163-176 192-198 231-240 1-125 I043F12 1344 143-250 164-174 190-196229-239 1-127 I043H07 1345 141-251 163-176 192-198 231-240 1-125 I044A111346 144-251 165-175 191-197 230-240 1-128 I044B11 1347 139-249 161-173189-195 228-238 1-123 I044C09 1348 140-250 162-174 190-196 229-239 1-124I044C10 1349 143-253 165-177 193-199 232-242 1-127 I044D03 1350 144-254166-179 195-201 234-243 1-128 I044D09 1351 141-251 163-176 192-198231-240 1-125 I044E07 1352 137-247 159-172 188-194 227-236 1-121 I044E111353 143-253 165-178 194-200 233-242 1-127 I044F07 1354 147-257 169-182198-204 237-246 1-131 I044G02 1355 141-251 163-176 192-198 231-240 1-125I044G07 1356 149-259 171-184 200-206 239-248 1-133 I044H01 1357 144-251165-175 191-197 230-240 1-128 I050A01 1358 142-253 164-177 193-199232-242 1-125 I050B12 1359 142-253 164-177 193-199 232-242 1-125 I0500061360 142-248 165-175 191-197 230-237 1-124 I050008 1361 142-253 164-177193-199 232-242 1-125 I050E01 1362 141-252 163-176 192-198 231-241 1-124I050E10 1363 138-248 160-172 188-194 227-237 1-121 I050H08 1364 142-253164-177 193-199 232-242 1-125 I051A04 1365 148-258 170-183 199-205238-247 1-131 I051A08 1366 142-252 164-176 192-198 231-241 1-125 I051Al21367 143-250 164-174 190-196 229-239 1-127 I051B08 1368 143-253 165-177193-199 232-242 1-126 I051C06 1369 136-246 158-170 186-192 225-235 1-119I051G12 1370 143-250 164-174 190-196 229-239 1-127 I055A05 1371 134-244156-169 185-191 224-233 1-117 I055A11 1372 134-244 156-169 185-191224-233 1-117 I061A03 1373 141-251 163-176 192-198 231-240 1-124 I061A041374 143-251 165-175 191-197 230-240 1-125 I061A08 1375 142-253 164-176192-198 233-242 1-124 I061A09 1376 142-252 164-176 192-198 231-241 1-124I061A10 1377 141-249 163-173 189-195 228-238 1-124 I061B07 1378 141-252163-176 192-198 231-241 1-124 I061B09 1379 143-253 165-178 194-200233-242 1-127 I061B12 1380 141-251 163-176 192-198 231-240 1-125 I061C121381 138-248 160-173 189-195 228-237 1-122 I061D01 1382 137-247 159-172188-194 227-236 1-121 I061D03 1383 141-251 163-176 192-198 231-240 1-125I061D04 1384 140-247 161-171 187-193 226-236 1-124 I061D07 1385 141-248164-174 190-196 229-237 1-125 I061D09 1386 141-248 162-172 188-194227-237 1-125 I061D10 1387 141-248 162-172 188-194 227-237 1-125 I061E011388 141-248 162-172 188-194 227-237 1-125 I061E05 1389 142-251 163-175191-197 230-240 1-126 I061E09 1390 141-251 163-176 192-198 231-240 1-125I061E12 1391 133-240 154-164 180-186 219-229 1-117 I061F01 1392 146-256168-181 197-203 236-245 1-130 I061F09 1393 139-246 160-170 186-192225-235 1-123 I061F10 1394 145-252 166-176 192-198 231-241 1-129 I061F111395 141-251 163-176 192-198 231-240 1-125 I061G01 1396 141-251 163-176192-198 231-240 1-125 I061G03 1397 141-251 163-176 192-198 231-240 1-125I061G09 1398 144-254 166-179 195-201 234-243 1-128 I061G10 1399 143-253165-178 194-200 233-242 1-127 I061G11 1400 137-247 159-171 187-193226-236 1-121 I061H05 1401 142-252 164-177 193-199 232-241 1-126 I064A051402 142-249 163-173 189-195 228-238 1-126 I064A11 1403 138-248 160-173189-195 228-237 1-122 I064B01 1404 138-248 160-173 189-195 228-237 1-122I064B02 1405 144-254 166-179 195-201 234-243 1-128 I064B12 1406 133-243155-168 184-190 223-232 1-117 I064C06 1407 145-255 167-180 196-202235-244 1-129 I064D01 1408 141-251 163-176 192-198 231-240 1-125 I064D021409 146-256 168-181 197-203 236-245 1-130 I064E01 1410 143-250 166-176192-198 231-239 1-127 I064E02 1411 140-251 162-174 190-196 229-240 1-124I064E03 1412 144-254 166-179 195-201 234-243 1-128 I064E07 1413 140-250162-175 191-197 230-239 1-124 I064E08 1414 140-250 162-174 190-196229-239 1-124 I064F05 1415 142-252 164-177 193-199 232-241 1-126 I064F081416 145-252 166-176 192-198 231-241 1-129 I064G06 1417 141-248 162-172188-194 227-237 1-125 I065Al2 1418 143-253 165-178 194-200 233-242 1-127I065C04 1419 139-249 161-173 189-195 228-238 1-123 I065C09 1420 140-250162-174 190-196 229-239 1-124 I065E02 1421 141-248 164-174 190-196229-237 1-125 I065E04 1422 135-245 157-169 185-191 224-234 1-119 I065F031423 137-247 159-172 188-194 227-236 1-121 I065G06 1424 135-242 156-166182-188 221-231 1-119 I065G07 1425 142-249 163-173 189-195 228-238 1-126I065G08 1426 139-246 160-170 186-192 225-235 1-123 I065H06 1427 144-254166-179 195-201 234-243 1-128 I066A03 1428 144-254 166-179 195-201234-243 1-128 I066A08 1429 137-247 159-171 187-193 226-236 1-121 I066A091430 135-245 157-169 185-191 224-234 1-119 I066A10 1431 142-252 164-177193-199 232-241 1-126 I066A11 1432 143-253 165-178 194-200 233-242 1-127I066B02 1433 135-242 156-166 182-188 221-231 1-119 I066B08 1434 137-247159-172 188-194 227-236 1-121 I066B10 1435 142-252 164-177 193-199232-241 1-126 I066C02 1436 135-245 157-169 185-191 224-234 1-119 I066C111437 141-251 163-176 192-198 231-240 1-125 I066C12 1438 135-242 156-166182-188 221-231 1-119 I066D06 1439 140-250 162-175 191-197 230-239 1-124I066D08 1440 138-248 160-173 189-195 228-237 1-122 I066D11 1441 144-254166-179 195-201 234-243 1-128 I066D12 1442 139-249 161-174 190-196229-238 1-123 I066E06 1443 137-247 159-171 187-193 226-236 1-121 I066E121444 135-242 156-166 182-188 221-231 1-119 I066G05 1445 142-249 163-173189-195 228-238 1-126 I066G08 1446 141-248 164-174 190-196 229-237 1-125I066G10 1447 144-254 166-178 194-200 233-243 1-128 I066G12 1448 143-254165-177 193-199 232-243 1-127 I066H04 1449 143-253 165-178 194-200233-242 1-127 I067A07 1450 144-254 166-179 195-201 234-243 1-128 I067A111451 135-245 157-170 186-192 225-234 1-119 I067B08 1452 149-259 171-184200-206 239-248 1-133 I067C08 1453 148-258 170-183 199-205 238-247 1-132I067C09 1454 143-253 165-178 194-200 233-242 1-127 I067D07 1455 137-247159-171 187-193 226-236 1-121 I067E01 1456 142-248 164-174 190-196229-238 1-124 I067E06 1457 135-245 157-169 185-191 224-234 1-119 I067E071458 150-260 172-184 200-206 239-249 1-134 I067E11 1459 141-251 163-176192-198 231-240 1-125 I067G03 1460 140-250 162-175 191-197 230-239 1-124I067G05 1461 140-250 162-174 190-196 229-239 1-124 I067G12 1462 141-252163-176 192-198 231-241 1-125 I067H05 1463 146-256 168-180 196-202235-245 1-130 I067H06 1464 135-245 157-169 185-191 224-234 1-119 I068C091465 138-248 160-172 188-194 227-237 1-121 I068G03 1466 144-254 166-178194-200 233-243 1-127 I068G04 1467 143-252 165-178 194-200 233-241 1-126I068G07 1468 142-251 164-174 190-196 229-240 1-124 I068G08 1469 144-254166-178 194-200 233-243 1-127 I070F07 1470 140-247 161-171 187-193226-236 1-124 I070G05 1471 140-250 162-175 191-197 230-239 1-124 I070H021472 141-248 162-172 188-194 227-237 1-125 I071A01 1473 141-251 163-176192-198 231-240 1-125 I071A03 1474 143-250 164-174 190-196 229-239 1-127I071B08 1475 144-252 166-176 192-198 231-241 1-126 I071E01 1476 138-248160-173 189-195 228-237 1-122 I071F11 1477 135-245 157-169 185-191224-234 1-119 I071G11 1478 141-251 163-176 192-198 231-240 1-125 I071H081479 141-251 163-176 192-198 231-240 1-125 I074A02 1480 142-250 164-174190-196 229-239 1-125 I074A08 1481 148-259 170-182 198-204 237-248 1-131I074D10 1482 146-253 168-178 194-200 233-242 1-128 I074E01 1483 146-255168-178 194-200 233-244 1-128 I074E02 1484 142-250 164-174 190-196229-239 1-124 I074E08 1485 144-259 166-179 195-205 240-248 1-127 I074F121486 142-250 164-174 190-196 229-239 1-124 I074H06 1487 140-251 162-175191-197 230-240 1-123 I074H07 1488 145-253 167-177 193-199 232-242 1-127I074H08 1489 143-254 165-178 194-200 233-243 1-126 I075A07 1490 145-253167-177 193-199 232-242 1-127 I075B01 1491 134-244 156-168 184-190223-233 1-117 I075B04 1492 134-247 156-169 185-191 224-236 1-117 I075B061493 141-252 163-175 191-197 230-241 1-124 I075B08 1494 144-257 166-179195-201 234-246 1-127 I075B09 1495 142-252 164-176 192-198 231-241 1-125I075B12 1496 141-251 163-176 192-198 231-240 1-124 I075C01 1497 148-259170-183 199-205 238-248 1-131 I075C05 1498 134-244 156-168 184-190223-233 1-117 I075D05 1499 145-253 168-179 195-201 234-242 1-127 I075D071500 142-252 164-176 192-198 231-241 1-125 I075D08 1501 141-251 163-175191-197 230-240 1-124 I075E01 1502 145-253 167-177 193-199 232-242 1-127I075E03 1503 150-261 172-184 200-206 239-250 1-132 I075E04 1504 144-255166-179 195-201 234-244 1-127 I075E05 1505 141-252 163-176 192-198231-241 1-124 I075E10 1506 141-252 163-176 192-198 231-241 1-124 I075E111507 134-244 156-168 184-190 223-233 1-117 I075E12 1508 143-254 165-178194-200 233-243 1-126 I075F02 1509 146-253 168-178 194-200 233-242 1-128I075F04 1510 142-251 164-176 192-198 231-240 1-125 I075F06 1511 146-254168-178 194-200 233-243 1-128 I075F07 1512 141-251 163-175 191-197230-240 1-124 I075F08 1513 134-244 156-168 184-190 223-233 1-117 I075F091514 147-257 169-181 197-203 236-246 1-129 I075F10 1515 135-243 157-167183-189 222-232 1-117 I075F11 1516 134-245 156-169 185-191 224-234 1-117I075G05 1517 141-252 163-175 191-197 230-241 1-124 I075G07 1518 141-252163-175 191-197 230-241 1-124 I075G08 1519 141-252 163-176 192-198231-241 1-124 I075G11 1520 142-253 164-177 193-199 232-242 1-125 I075G121521 134-245 156-169 185-191 224-234 1-117 I075H02 1522 144-254 166-178194-200 233-243 1-127 I075H03 1523 134-245 156-169 185-191 224-234 1-117I075H06 1524 134-244 156-168 184-190 223-233 1-117 I075H08 1525 144-254166-179 195-201 234-243 1-127 I076A01 1526 144-253 166-176 192-198231-242 1-126 I076A03 1527 137-247 159-171 187-193 226-236 1-119 I076A061528 134-245 156-168 184-190 223-234 1-117 I076A07 1529 140-250 162-174190-196 229-239 1-123 I076A08 1530 144-253 166-176 192-198 231-242 1-126I076B01 1531 145-257 167-179 195-201 236-246 1-127 I076B03 1532 134-245156-169 185-191 224-234 1-117 I076B07 1533 135-243 157-167 183-189222-232 1-117 I076B08 1534 143-252 166-177 193-199 232-241 1-125 I076C041535 142-250 164-174 190-196 229-239 1-124 I076C10 1536 141-251 163-175191-197 230-240 1-124 I076D01 1537 142-252 164-176 192-198 231-241 1-125I076D08 1538 141-251 163-175 191-197 230-240 1-124 I076D11 1539 144-255166-179 195-201 234-244 1-127 I076D12 1540 142-250 164-174 190-196229-239 1-124 I076E04 1541 145-252 167-177 193-199 232-241 1-127 I076E071542 141-251 163-175 191-197 230-240 1-124 I076E09 1543 142-253 164-177193-199 232-242 1-125 I076E11 1544 144-254 166-179 195-201 234-243 1-127I076F01 1545 144-253 166-178 194-199 232-242 1-127 I076F03 1546 141-251163-175 191-197 230-240 1-124 I076F04 1547 135-245 157-169 185-191224-234 1-117 I076F08 1548 142-250 164-174 190-196 229-239 1-124 I076F101549 141-252 163-175 191-197 230-241 1-124 I076G09 1550 134-245 156-168184-190 223-234 1-117 I076G10 1551 141-251 163-175 191-197 230-240 1-124I076G11 1552 144-259 166-179 195-205 240-248 1-127 I076G12 1553 147-257169-181 197-203 236-246 1-130 I076H02 1554 141-251 163-175 191-197230-240 1-124 I076H04 1555 143-251 165-175 191-197 230-240 1-125 I076H051556 141-251 163-175 191-197 230-240 1-124 I076H06 1557 141-252 163-176192-198 231-241 1-124 I076H09 1558 144-256 166-179 195-201 234-245 1-127I076H10 1559 144-256 166-179 195-201 234-245 1-127 I077D06 1560 140-250162-175 191-197 230-239 1-124 I078B04 1561 140-250 162-175 191-197230-239 1-124 I078E10 1562 141-251 163-176 192-198 231-240 1-125I002A01-K 1563 142-250 164-174 190-196 229-239 1-125 I002A01-R 1564142-250 164-174 190-196 229-239 1-125 I026C04-K 1565 142-250 164-176192-198 231-239 1-125 I026C04-R 1566 142-250 164-176 192-198 231-2391-125 I067B10 1567 149-259 171-183 199-205 238-248 1-133 I068C06 1568134-244 156-169 185-191 224-233 1-117 I075F12 1569 134-244 156-168184-190 223-233 1-117 I003C06 1570 141-249 163-173 189-195 228-238 1-124I025B06 1571 141-249 163-175 191-197 230-238 1-124 I025B09 1572 141-249163-175 191-197 230-238 1-124 I026C04 1573 141-249 163-175 191-197230-238 1-124 I027B12 1574 142-250 164-174 190-196 229-239 1-125 I030A101575 141-252 163-176 192-198 231-241 1-124 I064C04 1576 147-257 169-182198-204 237-246 1-131 I064C07 1577 134-241 157-167 183-189 222-230 1-118I065D04 1578 144-254 166-179 195-201 234-243 1-128 I065D08 1579 147-257169-182 198-204 237-246 1-131 I065F08 1580 135-242 158-168 184-190223-231 1-119 I067F05 1581 140-250 162-175 191-197 230-239 1-124 I068B041582 134-244 156-168 184-190 223-233 1-117 I068B08 1583 141-252 163-175191-197 231-241 1-124 I068C08 1584 143-254 165-178 194-200 233-243 1-126I068F03 1585 140-251 162-175 191-197 230-240 1-123 I069B07 1586 141-251163-176 192-198 231-240 1-125 I071B03 1587 141-251 163-176 192-198231-240 1-125 I072B09 1588 141-248 162-172 188-194 227-237 1-125 I073F041589 136-246 158-171 187-193 226-235 1-120 I074B12 1590 142-252 164-176192-198 231-241 1-124 I075A02 1591 141-251 163-175 191-197 230-240 1-124I075G01 1592 142-251 164-174 190-196 229-240 1-124 I078D02 1593 140-250162-175 191-197 230-239 1-124 I078D08 1594 144-251 165-175 191-197230-240 1-128 I078H08 1595 140-250 162-175 191-197 230-239 1-124 I064A031596 150-257 171-181 197-203 236-246 1-134 I064B03 1597 145-255 167-179195-201 234-244 1-129 I064B05 1598 140-250 162-174 190-196 229-239 1-124I064B11 1599 138-248 160-173 189-195 228-237 1-122 I064C02 1600 146-256168-180 196-202 235-245 1-130 I064C03 1601 140-250 162-175 191-197230-239 1-124 I064C11 1602 143-253 165-178 194-200 233-242 1-127 I064C121603 148-255 171-181 197-203 236-244 1-132 I064D03 1604 146-256 168-181197-203 236-245 1-130 I064D04 1605 141-251 163-176 192-198 231-240 1-125I064D06 1606 134-244 156-169 185-191 224-233 1-118 I064E05 1607 146-256168-180 196-202 235-245 1-130 I064E06 1608 145-255 167-180 196-202235-244 1-129 I064F07 1609 141-248 162-172 188-194 227-237 1-125 I064F091610 147-257 169-181 197-203 236-246 1-131 I064F10 1611 143-253 165-177193-199 232-242 1-127 I064F11 1612 142-252 164-177 193-199 232-241 1-126I064G01 1613 140-250 162-175 191-197 230-239 1-124 I064G04 1614 133-243155-167 183-189 222-232 1-117 I064G08 1615 138-245 159-169 185-191224-234 1-122 I064G10 1616 140-250 162-175 191-197 230-239 1-124 I064G111617 138-248 160-173 189-195 228-237 1-122 I064G12 1618 139-249 161-173189-195 228-238 1-123 I064H03 1619 143-253 165-178 194-200 233-242 1-127I064H04 1620 142-249 163-173 189-195 228-238 1-126 I064H06 1621 149-256170-180 196-202 235-245 1-133 I065A02 1622 141-248 162-172 188-194227-237 1-125 I065A04 1623 141-248 162-172 188-194 227-237 1-125 I065A061624 141-248 162-172 188-194 227-237 1-125 I065A07 1625 144-254 166-179195-201 234-243 1-128 I065B01 1626 145-255 167-180 196-202 235-244 1-129I065B05 1627 142-252 164-177 193-199 232-241 1-126 I065B09 1628 146-253167-177 193-199 232-242 1-130 I065B12 1629 139-249 161-174 190-196229-238 1-123 I065C02 1630 136-246 158-170 186-192 225-235 1-120 I065C061631 141-253 163-175 191-197 230-242 1-125 I065C08 1632 141-250 163-176192-198 231-239 1-125 I065C10 1633 137-247 159-172 188-194 227-236 1-121I065D01 1634 142-252 164-177 193-199 232-241 1-126 I065D03 1635 142-249165-175 191-197 230-238 1-126 I065D05 1636 143-253 165-178 194-200233-242 1-127 I065D06 1637 142-252 164-177 193-199 232-241 1-126 I065E011638 139-246 160-170 186-192 225-235 1-123 I065E05 1639 137-244 158-168184-190 223-233 1-121 I065E06 1640 146-256 168-181 197-203 236-245 1-130I065E08 1641 142-249 163-173 189-195 228-238 1-126 I065E09 1642 145-255167-179 195-201 234-244 1-129 I065E12 1643 141-248 162-172 188-194227-237 1-125 I065F04 1644 140-250 162-175 191-197 230-239 1-124 I065F051645 140-250 162-175 191-197 230-239 1-124 I065F07 1646 145-252 166-176192-198 231-241 1-129 I065F09 1647 143-250 164-174 190-196 229-239 1-127I065F12 1648 141-248 162-172 188-194 227-237 1-125 I065G01 1649 141-251163-176 192-198 231-240 1-125 I065G09 1650 143-253 165-178 194-200233-242 1-127 I065G10 1651 140-247 161-171 187-193 226-236 1-124 I065H051652 140-247 161-171 187-193 226-236 1-124 I065H07 1653 138-248 160-173189-195 228-237 1-122 I066A05 1654 137-247 159-172 188-194 227-236 1-121I066A06 1655 139-246 160-170 186-192 225-235 1-123 I066Al2 1656 142-252164-177 193-199 232-241 1-126 I066B05 1657 141-248 162-172 188-194227-237 1-125 I066B11 1658 142-252 164-177 193-199 232-241 1-126 I066C061659 144-254 166-178 194-200 233-243 1-128 I066C10 1660 141-248 162-172188-194 227-237 1-125 I066D02 1661 137-247 159-172 188-194 227-236 1-121I066D07 1662 141-248 162-172 188-194 227-237 1-125 I066E01 1663 137-247159-172 188-194 227-236 1-121 I066E03 1664 149-259 171-184 200-206239-248 1-133 I066E04 1665 141-248 162-172 188-194 227-237 1-125 I066E051666 142-252 164-177 193-199 232-241 1-126 I066E07 1667 141-248 162-172188-194 227-237 1-125 I066E09 1668 141-248 162-172 188-194 227-237 1-125I066F01 1669 141-251 163-176 192-198 231-240 1-125 I066F03 1670 141-248162-172 188-194 227-237 1-125 I066F04 1671 141-251 163-175 191-197230-240 1-125 I066F07 1672 143-253 165-178 194-200 233-242 1-127 I066F081673 144-254 166-179 195-201 234-243 1-128 I066F11 1674 142-252 164-177193-199 232-241 1-126 I066F12 1675 141-248 162-172 188-194 227-237 1-125I066G06 1676 143-250 164-174 190-196 229-239 1-127 I066G07 1677 133-243155-168 184-190 223-232 1-117 I066H02 1678 135-242 156-166 182-188221-231 1-119 I067A02 1679 141-248 162-172 188-194 227-237 1-125 I067A031680 137-247 159-172 188-194 227-236 1-121 I067A06 1681 141-248 162-172188-194 227-237 1-125 I067A08 1682 137-247 159-171 187-193 226-236 1-121I067A10 1683 140-250 162-175 191-197 230-239 1-124 I067B03 1684 142-253164-177 193-199 232-242 1-126 I067B04 1685 137-247 159-172 188-194227-236 1-121 I067C03 1686 134-244 156-169 185-191 224-233 1-117 I067C051687 137-247 159-172 188-194 227-236 1-121 I067C07 1688 141-251 163-176192-198 231-240 1-125 I067C10 1689 137-247 159-172 188-194 227-236 1-121I067C12 1690 142-252 164-177 193-199 232-241 1-126 I067D01 1691 136-246158-171 187-193 226-235 1-120 I067D03 1692 137-244 158-168 184-190223-233 1-121 I067D05 1693 146-256 168-180 196-202 235-245 1-130 I067D061694 137-244 158-168 184-190 223-233 1-121 I067D09 1695 142-252 164-177193-199 232-241 1-126 I067D12 1696 137-247 159-172 188-194 227-236 1-121I067E02 1697 137-247 159-172 188-194 227-236 1-121 I067E04 1698 142-252164-176 192-198 231-241 1-126 I067E05 1699 144-254 166-179 195-201234-243 1-128 I067F01 1700 141-248 164-174 190-196 229-237 1-125 I067F031701 144-254 166-179 195-201 234-243 1-128 I067F04 1702 139-246 160-170186-192 225-235 1-123 I067F08 1703 140-247 161-171 187-193 226-236 1-124I067F10 1704 137-247 159-172 188-194 227-236 1-121 I067F11 1705 140-248161-171 187-193 226-237 1-124 I067G01 1706 141-251 163-176 192-198231-240 1-125 I067G09 1707 137-247 159-171 187-193 226-236 1-121 I067H071708 144-251 165-175 191-197 230-240 1-128 I068A07 1709 143-254 165-178194-200 233-243 1-126 I068E05 1710 148-257 170-183 199-205 238-246 1-131I068E08 1711 135-247 157-169 185-193 226-236 1-117 I068E11 1712 141-251163-176 192-198 231-240 1-124 I068F04 1713 142-252 164-176 192-198231-241 1-125 I068G05 1714 137-245 159-169 185-191 224-234 1-119 I068G061715 140-250 162-174 190-196 229-239 1-123 I068G11 1716 147-258 169-182198-204 237-247 1-130 I069A09 1717 141-248 164-174 190-196 229-237 1-125I069A10 1718 141-248 162-172 188-194 227-237 1-125 I069B06 1719 141-248164-174 190-196 229-237 1-125 I069B09 1720 139-249 161-174 190-196229-238 1-123 I069B12 1721 141-248 162-172 188-194 227-237 1-125 I069C061722 143-250 164-174 190-196 229-239 1-127 I069C09 1723 143-250 164-174190-196 229-239 1-127 I069D03 1724 142-249 163-173 189-195 228-238 1-126I069E09 1725 142-249 163-173 189-195 228-238 1-126 I069E11 1726 140-247161-171 187-193 226-236 1-124 I069F05 1727 141-248 162-172 188-194227-237 1-125 I069F07 1728 141-248 162-172 188-194 227-237 1-125 I069F121729 140-247 161-171 187-193 226-236 1-124 I069G06 1730 142-249 163-173189-195 228-238 1-126 I069G08 1731 145-252 166-176 192-198 231-241 1-129I069G11 1732 141-248 162-172 188-194 227-237 1-125 I070A03 1733 141-248164-174 190-196 229-237 1-125 I070A09 1734 141-248 162-172 188-194227-237 1-125 I070B01 1735 144-254 166-179 195-201 234-243 1-128 I070B051736 141-251 163-176 192-198 231-240 1-125 I070D03 1737 141-248 164-174190-196 229-237 1-125 I070D04 1738 141-251 163-176 192-198 231-240 1-125I070E01 1739 144-254 166-179 195-201 234-243 1-128 I070F01 1740 144-251165-175 191-197 230-240 1-128 I070G10 1741 141-248 162-172 188-194227-237 1-125 I071A06 1742 135-242 156-166 182-188 221-231 1-119 I071B021743 135-245 157-170 186-192 225-234 1-119 I071D02 1744 137-247 159-172188-194 227-236 1-121 I071D08 1745 146-256 168-181 197-203 236-245 1-130I071F01 1746 137-247 159-172 188-194 227-236 1-121 I071G09 1747 141-251163-176 192-198 231-240 1-125 I072A01 1748 139-249 161-174 190-196229-238 1-123 I072A09 1749 141-251 163-176 192-198 231-240 1-125 I072B021750 135-245 157-170 186-192 225-234 1-119 I072B10 1751 137-247 159-172188-194 227-236 1-121 I072B11 1752 141-251 163-176 192-198 231-240 1-125I072B12 1753 140-249 162-173 189-195 228-238 1-124 I072C05 1754 135-245157-169 185-191 224-234 1-119 I072C10 1755 141-248 162-172 188-194227-237 1-125 I072D01 1756 141-251 163-176 192-198 231-240 1-125 I072D051757 135-245 157-169 185-191 224-234 1-119 I072E01 1758 141-251 163-176192-198 231-240 1-125 I072E04 1759 144-254 166-179 195-201 234-243 1-128I072E05 1760 141-251 163-176 192-198 231-240 1-125 I072E06 1761 135-242156-166 182-188 221-231 1-119 I072F03 1762 135-242 156-166 182-188221-231 1-119 I072F07 1763 141-251 163-176 192-198 231-240 1-125 I072F111764 140-247 161-171 187-193 226-236 1-124 I072G03 1765 141-248 162-172188-194 227-237 1-125 I072G04 1766 137-247 159-171 187-193 226-236 1-121I072G05 1767 137-247 159-171 187-193 226-236 1-121 I072G09 1768 141-251163-176 192-198 231-240 1-125 I072H03 1769 141-251 163-176 192-198231-240 1-125 I072H07 1770 137-247 159-172 188-194 227-236 1-121 I073A021771 141-248 164-174 190-196 229-237 1-125 I073A03 1772 142-252 164-177193-199 232-241 1-126 I073A04 1773 148-258 170-183 199-205 238-247 1-132I073A05 1774 141-251 163-176 192-198 231-240 1-125 I073A06 1775 141-251163-176 192-198 231-240 1-125 I073A09 1776 141-251 163-176 192-198231-240 1-125 I073A10 1777 146-253 167-177 193-199 232-242 1-130 I073A111778 141-248 164-174 190-196 229-237 1-125 I073B02 1779 144-254 166-179195-201 234-243 1-128 I073B05 1780 144-254 166-179 195-201 234-243 1-128I073B06 1781 139-246 160-170 186-192 225-235 1-123 I073B07 1782 138-248160-173 189-195 228-237 1-122 I073B08 1783 141-251 163-176 192-198231-240 1-125 I073B11 1784 141-251 163-176 192-198 231-240 1-125 I073C011785 141-248 162-172 188-194 227-237 1-125 I073C02 1786 148-255 169-179195-201 234-244 1-132 I073C04 1787 142-252 164-177 193-199 232-241 1-125I073C07 1788 134-241 155-165 181-187 220-230 1-118 I073C08 1789 142-252164-177 193-199 232-241 1-126 I073C09 1790 141-248 162-172 188-194227-237 1-125 I073C11 1791 146-256 168-181 197-203 236-245 1-130 I073C121792 141-251 163-176 192-198 231-240 1-125 I073D01 1793 141-251 163-176192-198 231-240 1-125 I073D03 1794 135-245 157-169 185-191 224-234 1-119I073D06 1795 141-251 163-176 192-198 231-240 1-125 I073D08 1796 144-254166-179 195-201 234-243 1-128 I073D10 1797 140-250 162-175 191-197230-239 1-124 I073D11 1798 141-251 163-176 192-198 231-240 1-125 I073E011799 148-258 170-183 199-205 238-247 1-132 I073E02 1800 141-251 163-176192-198 231-240 1-125 I073E03 1801 141-251 163-176 192-198 231-240 1-125I073E05 1802 141-251 163-176 192-198 231-240 1-125 I073E06 1803 141-251163-176 192-198 231-240 1-125 I073E08 1804 140-250 162-175 191-197230-239 1-124 I073F01 1805 141-251 163-175 191-197 230-240 1-125 I073F021806 141-251 163-175 191-197 230-240 1-125 I073F03 1807 141-251 163-176192-198 231-240 1-125 I073F05 1808 141-251 163-176 192-198 231-240 1-125I073F07 1809 141-251 163-175 191-197 230-240 1-125 I073F09 1810 141-251163-176 192-198 231-240 1-125 I073F11 1811 141-251 163-176 192-198231-240 1-125 I073F12 1812 141-251 163-175 191-197 230-240 1-125 I073G031813 143-253 165-178 194-200 233-242 1-127 I073G04 1814 143-253 165-178194-200 233-242 1-127 I073G05 1815 135-245 157-169 185-191 224-234 1-119I073G06 1816 141-248 162-172 188-194 227-237 1-125 I073G07 1817 142-249163-173 189-195 228-238 1-126 I073G08 1818 139-246 160-170 186-192225-235 1-123 I073G09 1819 145-255 167-180 196-202 235-244 1-129 I073G101820 135-245 157-170 186-192 225-234 1-119 I073G12 1821 142-252 164-177193-199 232-241 1-126 I073H01 1822 141-251 163-176 192-198 231-240 1-125I073H03 1823 141-248 162-172 188-194 227-237 1-125 I073H05 1824 141-251163-176 192-198 231-240 1-125 I073H06 1825 141-251 163-176 192-198231-240 1-125 I073H07 1826 138-245 159-169 185-191 224-234 1-122 I073H081827 141-251 163-176 192-198 231-240 1-125 I074A05 1828 144-255 166-179195-201 234-244 1-127 I074A06 1829 145-253 167-177 193-199 232-242 1-127I074B03 1830 134-242 156-166 182-188 221-231 1-117 I074B11 1831 140-251162-175 191-197 230-240 1-123 I074C07 1832 141-251 163-175 191-197230-240 1-124 I074D03 1833 143-251 165-175 191-197 230-240 1-125 I074D041834 134-246 156-169 185-191 224-235 1-117 I074D05 1835 145-253 167-177193-199 232-242 1-127 I074D07 1836 151-262 173-186 202-208 241-251 1-134I074D08 1837 141-251 163-175 191-197 230-240 1-124 I074D11 1838 139-249161-174 190-196 229-238 1-122 I074E05 1839 134-245 156-169 185-191224-234 1-117 I074E07 1840 141-251 163-175 191-197 230-240 1-124 I074E091841 147-258 169-182 198-204 237-247 1-130 I074E11 1842 138-244 160-170186-192 225-233 1-121 I074H05 1843 144-254 166-178 194-200 233-243 1-126I075A03 1844 135-242 158-168 184-190 223-231 1-117 I075A10 1845 135-244157-169 185-191 224-233 1-117 I075B07 1846 144-254 166-178 194-200233-243 1-127 I075D11 1847 134-246 156-169 185-191 224-235 1-117 I075D121848 145-253 167-177 193-199 232-242 1-127 I075G02 1849 144-255 166-179195-201 234-244 1-127 I075G09 1850 143-253 165-177 193-199 232-242 1-126I075G10 1851 140-250 162-174 190-196 229-239 1-122 I075H05 1852 142-252164-176 192-198 231-241 1-125 I075H07 1853 145-253 167-177 193-199232-242 1-127 I076A11 1854 142-254 164-177 193-199 232-243 1-125 I076Al21855 144-256 166-178 194-200 233-245 1-127 I076B06 1856 142-249 164-174190-196 229-238 1-124 I076B10 1857 142-254 164-177 193-199 232-243 1-125I076B12 1858 145-253 167-177 193-199 232-242 1-127 I076C06 1859 143-253165-177 193-199 232-242 1-126 I076C11 1860 134-245 156-168 184-190223-234 1-117 I076D06 1861 141-252 163-176 192-198 231-241 1-124 I076E051862 144-255 166-179 195-201 234-244 1-127 I076E08 1863 135-243 157-167183-189 222-232 1-117 I076F06 1864 134-245 156-169 185-191 224-234 1-117I076G01 1865 144-254 166-178 194-200 233-243 1-127 I076H01 1866 146-254168-178 194-200 233-243 1-128 I076H03 1867 142-250 164-174 190-196229-239 1-124 I077B05 1868 147-257 169-182 198-204 237-246 1-131 I077C101869 141-251 163-176 192-198 231-240 1-125 I077D01 1870 141-251 163-176192-198 231-240 1-125 I077D04 1871 141-248 162-172 188-194 227-237 1-125I077D11 1872 141-251 163-176 192-198 231-240 1-125 I077D12 1873 140-247161-171 187-193 226-236 1-124 I077E01 1874 142-252 164-177 193-199232-241 1-126 I077E03 1875 142-252 164-177 193-199 232-241 1-126 I077E081876 141-248 164-174 190-196 229-237 1-125 I077F05 1877 141-248 162-172188-194 227-237 1-125 I077G06 1878 141-251 163-176 192-198 231-240 1-125I077H02 1879 141-248 164-174 190-196 229-237 1-125 I078B05 1880 143-253165-178 194-200 233-242 1-127 I079E02 1881 137-244 160-170 186-192225-233 1-121 I079F11 1882 132-239 155-165 181-187 220-228 1-116 I082G021883 136-243 159-169 185-191 224-232 1-120 I082H08 1884 132-242 154-167183-189 222-231 1-115 I099D03 1885 137-247 159-172 188-194 227-236 1-120I079B05 1886 130-240 152-165 181-187 220-229 1-114 I079B12 1887 134-241157-167 183-189 222-230 1-118 I079C01 1888 131-241 153-166 182-188221-230 1-115 I079F06 1889 134-241 157-167 183-189 222-230 1-118 I079F081890 138-248 160-172 188-194 227-237 1-122 I080A03 1891 139-249 161-173189-195 228-238 1-122 I080A08 1892 136-247 158-171 187-193 226-236 1-119I080B01 1893 144-254 166-178 194-200 233-243 1-126 I080D03 1894 139-249161-173 189-195 228-238 1-122 I080E05 1895 142-253 164-177 193-199232-242 1-125 I080G07 1896 138-245 161-172 188-194 227-234 1-120 I080G091897 137-249 159-172 188-194 227-238 1-120 I082A05 1898 131-240 153-165181-187 220-229 1-115 I082B08 1899 137-247 159-171 187-193 226-236 1-121I082C03 1900 138-245 161-171 187-193 226-234 1-122 I082D07 1901 134-241157-167 183-189 222-230 1-118 I082G01 1902 138-245 161-171 187-193226-234 1-122 I083B12 1903 139-247 161-171 187-193 226-236 1-121 I083G031904 139-249 161-173 189-195 228-238 1-122 I084A01 1905 130-240 152-164180-186 219-229 1-114 I084B02 1906 130-237 153-163 179-185 218-226 1-114I084C04 1907 131-238 152-162 178-184 217-227 1-115 I084C11 1908 134-244156-169 185-191 224-233 1-118 I079A01 1909 134-243 156-168 184-190223-232 1-118 I079A03 1910 134-244 156-169 185-191 224-233 1-118 I079A041911 134-241 155-165 181-187 220-230 1-118 I079A06 1912 133-240 154-164180-186 219-229 1-117 I079A07 1913 136-246 158-170 186-192 225-235 1-120I079A10 1914 148-255 169-179 195-201 234-244 1-132 I079A11 1915 135-242158-168 184-190 223-231 1-119 I079B02 1916 134-243 156-168 184-190223-232 1-118 I079B03 1917 136-246 158-170 186-192 225-235 1-120 I079B041918 130-240 152-165 181-187 220-229 1-114 I079B07 1919 138-245 159-169185-191 224-234 1-122 I079B09 1920 139-246 162-172 188-194 227-235 1-123I079C02 1921 144-251 167-177 193-199 232-240 1-128 I079C04 1922 132-239155-165 181-187 220-228 1-116 I079C05 1923 140-247 163-173 189-185228-236 1-124 I079C07 1924 137-244 158-168 184-190 223-233 1-121 I079D011925 144-254 166-179 195-201 234-243 1-128 I079D02 1926 135-245 157-169185-191 224-234 1-119 I079D04 1927 133-243 155-167 183-189 222-232 1-117I079D06 1928 137-247 159-171 187-193 226-236 1-121 I079D07 1929 136-243157-167 183-189 222-232 1-120 I079D08 1930 130-240 152-165 181-187220-229 1-114 I079D09 1931 131-238 152-162 178-184 217-227 1-115 I079D111932 134-241 157-167 183-189 222-230 1-118 I079E06 1933 136-244 158-168184-190 223-233 1-120 I079E08 1934 137-247 159-171 187-193 226-236 1-121I079E11 1935 136-243 159-169 185-191 224-232 1-120 I079E12 1936 143-253165-177 193-199 232-242 1-127 I079F01 1937 133-241 154-164 180-186219-230 1-117 I079F02 1938 148-253 169-179 195-201 234-242 1-132 I079F031939 140-247 161-171 187-193 226-236 1-124 I079F04 1940 136-243 157-167183-189 222-232 1-120 I079F09 1941 130-237 151-161 177-183 216-226 1-114I079F10 1942 136-243 157-167 183-189 222-232 1-120 I079F12 1943 136-243159-169 185-191 224-232 1-120 I079G02 1944 136-243 157-167 183-189222-232 1-120 I079G05 1945 136-243 159-169 185-191 224-232 1-120 I079G061946 135-245 157-170 186-192 225-234 1-119 I079H05 1947 136-243 157-167183-189 222-232 1-120 I079H06 1948 134-241 157-167 183-189 222-230 1-118I080A01 1949 132-242 154-166 182-188 221-231 1-115 I080A02 1950 134-245156-169 185-191 224-234 1-117 I080A05 1951 142-253 164-177 193-199232-242 1-125 I080A06 1952 143-250 166-176 192-198 231-239 1-125 I080A071953 136-247 158-171 187-193 226-236 1-119 I080A10 1954 142-252 164-176192-198 231-241 1-125 I080B02 1955 140-248 162-172 187-193 227-237 1-122I080B03 1956 139-249 161-173 189-195 228-238 1-122 I080B05 1957 139-249161-173 189-195 228-238 1-121 I080B06 1958 143-254 165-177 193-199232-243 1-126 1080B07 1959 135-243 157-167 183-189 222-232 1-117 I080B081960 137-248 159-172 188-194 227-237 1-120 I080B09 1961 143-254 165-178194-200 233-243 1-126 I080B10 1962 139-249 161-173 189-195 228-238 1-122I080B11 1963 138-248 160-172 188-194 227-237 1-121 I080B12 1964 141-253164-179 195-201 234-242 1-123 I080003 1965 140-248 162-172 188-194227-237 1-122 I080006 1966 146-254 168-178 194-200 233-243 1-128 I0800071967 145-256 167-180 196-202 235-245 1-128 I080008 1968 138-249 160-173189-195 228-238 1-121 I080C10 1969 132-243 154-167 183-189 222-232 1-115I080C11 1970 138-249 160-173 189-195 228-238 1-121 I080C12 1971 139-249161-173 189-195 228-238 1-122 I080D01 1972 138-245 161-171 187-193226-234 1-120 I080D02 1973 142-254 164-177 193-199 232-243 1-125 I080D041974 140-248 162-172 188-194 227-237 1-122 I080D05 1975 138-246 160-170186-192 225-235 1-120 I080D08 1976 138-248 160-172 188-194 227-237 1-121I080D09 1977 139-250 161-174 190-196 229-239 1-122 I080D11 1978 136-247158-171 187-193 226-236 1-119 I080D12 1979 137-245 159-169 185-191224-234 1-119 I080E01 1980 138-246 160-170 186-192 225-235 1-120 I080E041981 137-247 159-171 187-193 226-236 1-120 I080E06 1982 138-248 160-172188-194 227-237 1-121 I080E07 1983 143-254 165-178 194-200 233-243 1-126I080E08 1984 138-249 160-173 189-195 228-238 1-121 I080E12 1985 132-242154-166 182-188 221-231 1-114 I080F04 1986 139-249 161-173 189-195228-238 1-122 I080F05 1987 143-253 165-177 193-199 232-242 1-126 I080F061988 140-248 162-172 188-194 227-237 1-122 I080F08 1989 132-240 154-164180-186 219-229 1-114 I080G03 1990 142-250 164-174 190-196 229-239 1-124I080G04 1991 133-244 156-171 187-193 226-233 1-115 I080G10 1992 145-252167-177 193-199 232-241 1-127 I080G11 1993 137-247 159-171 187-193226-236 1-120 I080H01 1994 142-252 164-176 192-198 231-241 1-124 I080H021995 140-248 162-172 188-194 227-237 1-123 I080H03 1996 136-246 158-170186-192 225-235 1-119 I080H04 1997 138-249 160-173 189-195 228-238 1-121I080H05 1998 137-247 159-171 187-193 226-236 1-120 I080H06 1999 138-249160-173 189-195 228-238 1-121 I080H07 2000 138-248 160-172 188-194227-237 1-121 I080H08 2001 140-251 162-175 191-197 230-240 1-122 I080H092002 141-249 163-173 189-195 228-238 1-123 I081A01 2003 130-237 153-163179-185 218-226 1-114 I081A03 2004 135-245 157-170 186-192 225-234 1-119I081A04 2005 130-237 153-163 179-185 218-226 1-114 I081A06 2006 130-237151-161 177-183 216-226 1-114 I081A08 2007 130-240 152-164 180-186219-229 1-114 I081A09 2008 134-241 155-165 181-187 220-230 1-118 I081A102009 133-243 155-168 184-190 223-232 1-117 I081B01 2010 130-236 151-161177-183 216-226 1-114 I081B04 2011 134-244 156-169 185-191 224-233 1-118I081B05 2012 133-243 155-168 184-190 223-232 1-117 I081B06 2013 133-240154-164 180-186 219-229 1-117 I081B07 2014 136-243 157-167 183-189222-232 1-120 I081B08 2015 132-239 153-163 179-185 218-228 1-116 I081B092016 130-240 152-164 180-186 219-229 1-114 I081B10 2017 130-237 153-163179-185 218-226 1-114 I081B11 2018 132-239 153-163 179-185 218-228 1-116I081C07 2019 130-237 153-163 179-185 218-226 1-114 I081C08 2020 130-237153-163 179-185 218-226 1-114 I081D04 2021 135-242 156-166 182-188221-231 1-119 I081D06 2022 132-239 153-163 179-185 218-228 1-116 I081D082023 132-239 153-163 179-185 218-228 1-116 I081D09 2024 130-238 152-162178-184 217-227 1-114 I081D10 2025 130-240 152-164 180-186 219-229 1-114I081D11 2026 134-244 156-169 185-191 224-233 1-118 I081D12 2027 130-237153-163 179-185 218-226 1-114 I081E02 2028 130-237 153-163 179-185218-226 1-114 I081E03 2029 130-240 152-164 180-186 219-229 1-114 I081E052030 130-240 152-164 180-186 219-229 1-114 I081E06 2031 134-241 155-165181-187 220-230 1-118 I081E07 2032 134-241 155-165 181-187 220-230 1-118I081E10 2033 142-249 163-173 189-195 228-238 1-126 I081F01 2034 130-239152-164 180-186 219-228 1-114 I081F04 2035 132-239 153-163 179-185218-228 1-116 I081F05 2036 130-237 151-161 177-183 216-226 1-114 I081F062037 134-244 156-169 185-191 224-233 1-118 I081F07 2038 132-239 153-163179-185 218-228 1-116 I081F11 2039 130-237 151-161 177-183 216-226 1-114I081G01 2040 130-237 153-163 179-185 218-226 1-114 I081G04 2041 130-240152-164 180-186 219-229 1-114 I081G06 2042 135-245 157-170 186-192225-234 1-119 I081G10 2043 130-237 153-163 179-185 218-226 1-114 I081H022044 130-240 152-164 180-186 219-229 1-114 I081H03 2045 130-240 152-164180-186 219-229 1-114 I081H04 2046 135-242 156-166 182-188 221-231 1-119I081H06 2047 130-240 152-165 181-187 220-229 1-114 I081H08 2048 130-240152-164 180-186 219-229 1-114 I082A02 2049 139-249 161-173 189-195228-238 1-123 I082A04 2050 130-240 152-165 181-187 220-229 1-114 I082A082051 134-243 156-168 184-190 223-232 1-118 I082A11 2052 130-240 152-165181-187 220-229 1-114 I082B06 2053 131-238 154-164 180-186 219-227 1-115I082B09 2054 134-241 157-167 183-189 222-230 1-118 I082B12 2055 131-241153-166 182-188 221-230 1-115 I082C01 2056 136-243 157-167 183-189222-232 1-120 I082C05 2057 136-243 157-167 183-189 222-232 1-120 I082C082058 137-244 158-168 184-190 223-233 1-121 I082D02 2059 130-240 152-165181-187 220-229 1-114 I082E05 2060 134-241 155-165 181-187 220-230 1-118I082E06 2061 130-240 152-165 181-187 220-229 1-114 I082E07 2062 139-246162-172 188-194 227-235 1-123 I082F11 2063 136-243 159-169 185-191224-232 1-120 I082G07 2064 136-243 159-169 185-191 224-232 1-120 I082G102065 138-249 160-173 189-195 228-238 1-118 I082G11 2066 143-250 164-174190-196 229-239 1-127 I082H04 2067 132-238 153-163 179-185 218-227 1-116I082H09 2068 139-246 160-170 186-192 225-235 1-123 I083A06 2069 137-244159-169 185-191 224-233 1-120 I083A09 2070 138-248 160-172 188-194227-237 1-121 I083A11 2071 136-248 158-171 187-193 226-237 1-119 I083B032072 139-247 161-171 187-193 226-236 1-121 I083B05 2073 139-250 161-174190-196 229-239 1-122 I083B06 2074 139-250 161-174 190-196 229-239 1-122I083B10 2075 139-246 162-172 188-194 227-235 1-121 I083C01 2076 136-247158-171 187-193 226-236 1-119 I083C02 2077 136-246 158-171 187-193226-235 1-119 I083C07 2078 137-249 159-172 188-194 227-238 1-120 I083C122079 136-246 158-171 187-193 226-235 1-119 I083D04 2080 146-256 168-181197-203 236-245 1-129 I083D07 2081 150-262 173-188 204-210 243-251 1-132I083D08 2082 143-254 165-178 194-200 233-243 1-126 I083D10 2083 147-258169-181 197-203 236-247 1-130 I083D12 2084 134-242 156-166 182-188221-231 1-116 I083E02 2085 139-249 161-173 189-195 228-238 1-122 I083E032086 136-248 158-171 187-193 226-237 1-119 I083E04 2087 144-255 166-179195-201 234-244 1-127 I083E08 2088 140-248 162-172 188-194 227-237 1-122I083E12 2089 135-245 157-170 186-192 225-234 1-118 I083F02 2090 146-258168-181 197-203 236-247 1-129 I083F04 2091 138-248 160-172 188-194227-237 1-121 I083F06 2092 135-247 157-170 186-192 225-236 1-118 I083F082093 139-250 161-174 190-196 229-239 1-122 I083F11 2094 137-248 159-172188-194 227-237 1-120 I083G04 2095 139-250 161-174 190-196 229-239 1-122I083G05 2096 139-249 161-173 189-195 228-238 1-121 I083G06 2097 139-250161-174 190-196 229-239 1-122 I083G08 2098 142-253 164-177 193-199232-242 1-125 I083G09 2099 132-242 154-166 182-188 221-231 1-114 I083G112100 141-252 163-176 192-198 231-241 1-124 I083H04 2101 142-253 164-177193-199 232-242 1-125 I083H05 2102 135-243 157-167 183-189 222-232 1-117I083H07 2103 139-247 161-171 187-193 226-236 1-121 I084A03 2104 130-237153-163 179-185 218-226 1-114 I084A08 2105 130-240 152-164 180-186219-229 1-114 I084B08 2106 135-242 156-166 182-188 221-231 1-119 I084C022107 136-243 157-167 183-189 222-232 1-120 I084D03 2108 130-240 152-164180-186 219-229 1-114 I084D05 2109 133-243 155-168 184-190 223-232 1-117I084E01 2110 130-237 153-163 179-185 218-226 1-114 I084E06 2111 130-237153-163 179-185 218-226 1-114 I084E10 2112 130-237 151-161 177-183216-226 1-114 I084E12 2113 130-240 152-164 180-186 219-229 1-114 I084F042114 130-237 153-163 179-185 218-226 1-114 I084F07 2115 130-237 153-163179-185 218-226 1-114 I084F12 2116 135-245 157-170 186-192 225-234 1-119I084G12 2117 130-240 152-164 180-186 219-229 1-114 I084H02 2118 130-237153-164 179-185 218-226 1-114 I099B05 2119 146-256 168-180 196-202235-245 1-129 I099G09 2120 139-249 161-173 189-195 228-238 1-122 I099H012121 140-248 162-172 188-194 227-237 1-122 I099H06 2122 139-249 161-173189-195 228-238 1-122 I099H08 2123 145-255 167-179 195-201 234-244 1-128I100A01 2124 137-247 159-172 188-194 227-236 1-120 I100A10 2125 141-251163-175 191-197 230-240 1-124 I100B03 2126 137-247 159-172 188-194227-236 1-120 I100B04 2127 137-247 159-172 188-194 227-236 1-120 I1000032128 141-251 163-175 191-197 230-240 1-124 BAB2001 3240 140-247 163-173189-195 228-236 1-124 BAB2080 3241 135-245 157-169 185-191 224-234 1-119BAB2015 3242 140-247 163-173 189-195 228-236 1-124 BAB2019 3243 141-251163-176 192-198 231-240 1-125 BAB2087 3244 144-253 167-177 193-199232-242 1-128 BAB2016 3245 141-251 163-176 192-198 231-246 1-125 BAB20343246 146-258 170-183 199-205 238-247 1-132 BAB2065 3247 141-250 162-172191-197 230-239 1-125 AAs of AAs of AAs of Clone ID VH CDR1 VH CDR2VH CDR3 Sequence (SEQ ID NO) I003F12S 26-35 50-66  99-111 HDDDVLTRGYYFES(SEQ ID NO: 2130) I006D08 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I008A11 26-37 52-69 102-117 DRYDILTGYYYYGMDV(SEQ ID NO: 2129) I017D10 26-35 50-66  99-121 VQMDSEYYDLLTGINVGPYYFDY(SEQ ID NO: 2132) I022D01 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I031F02 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I050A12 26-35 50-66  99-113 APYDLLTHYFHYFDY(SEQ ID NO: 2134) I051C04 26-35 50-66  99-118 AATTSQKHNKYAYYFYGMDV(SEQ ID NO: 2131) I050B11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I050B11-01 26-35 50-66  99-114 PFYDTLTSYVFQVWVA(SEQ ID NO: 2143) I050B11-02 26-35 50-66  99-114 PFYDTLTSYVFQVWVA(SEQ ID NO: 2143) I050B11-03 26-35 50-66  99-114 PFYDTLTRYVFQYFDH(SEQ ID NO: 2144) I050B11-04 26-35 50-66  99-114 PFYDTLTGYVFQYFDH(SEQ ID NO: 2141) I050B11-05 26-35 50-66  99-114 PFYDTLTRYVFQVWVA(SEQ ID NO: 2142) I050B11-06 26-35 50-66  99-114 PFYDTLTGYVFQVWVA(SEQ ID NO: 2140) I050B11-07 26-35 50-66  99-114 PFYDTLTRYVFQYFDH(SEQ ID NO: 2144) I050B11-08 26-35 50-66  99-114 PFYDTLTGYVFQYFDH(SEQ ID NO: 2141) I050B11-09 26-35 50-66  99-114 PFYDTLTRYVFQVWVA(SEQ ID NO: 2142) I050B11-10 26-35 50-66  99-114 PFYDTLTRYVFQVWVA(SEQ ID NO: 2142) I050B11-11 26-35 50-66  99-114 PFYDTLTGYVFQVWVA(SEQ ID NO: 2140) I050B11-12 26-35 50-66  99-114 PFYDTLTGYVFQVWVA(SEQ ID NO: 2140) I050B11-13 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I050B11-14 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I050B11-15 26-35 50-66  99-114 PFYDTLTSYVFQVWVA(SEQ ID NO: 2143) I050B11-16 26-35 50-66  99-114 PFYDTLTSYVFQVWVA(SEQ ID NO: 2143) I050B11-17 26-35 50-66  99-114 PFYDTLTRYVFQYFDH(SEQ ID NO: 2144) I050B11-18 26-35 50-66  99-114 PFYDTLTRYVFQYFDH(SEQ ID NO: 2144) I050B11-19 26-35 50-66  99-114 PFYDILTSYVFQYFDH(SEQ ID NO: 2139) I050B11-20 26-35 50-66  99-114 PFYDILTSYVFQYFDH(SEQ ID NO: 2139) I050B11-21 26-35 50-66  99-114 PFYDILTRYVFQYFDH(SEQ ID NO: 2138) I050B11-22 26-35 50-66  99-114 PFYDILTRYVFQYFDH(SEQ ID NO: 2138) I050B11-23 26-35 50-66  99-114 PFYDILTRYVFQYFDH(SEQ ID NO: 2138) I050B11-24 26-35 50-66  99-114 PFYDILTSYVFQYFDH(SEQ ID NO: 2139) I050B11-25 26-35 50-66  99-114 PFYDTLTRYVFQYFDH(SEQ ID NO: 2144) I050B11-26 26-35 50-66  99-114 PFYDILTSYVFQYFDH(SEQ ID NO: 2139) I050B11-27 26-35 50-66  99-114 PFYDILTRYVFQYFDH(SEQ ID NO: 2138) I050B11-28 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093D03 26-35 50-66  99-114 PFYDTLTSYVLGYYLS(SEQ ID NO: 2145) I093D09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093G08 26-35 50-66  99-114 PFYDTLTSYVFQVWVA(SEQ ID NO: 2143) I097D11 26-35 50-66  99-114 PFYDILTSYVFQYFDH(SEQ ID NO: 2139) I101A04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I101B01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I102A02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I102E01 26-35 50-66  99-114 PFYDTLTRYVFQYFDH(SEQ ID NO: 2144) I102G06 26-35 50-66  99-114 PFYDTLTGYVFQYFDH(SEQ ID NO: 2141) I087A07 26-35 50-66  99-114 PFYDTLTSYVLPRVIP(SEQ ID NO: 2227) I087A08 26-35 50-66  99-113 PFYDTLTSYVCRPHF(SEQ ID NO: 2238) I087A09 26-35 50-66  99-113 PFYDTLTSYVRCPYV(SEQ ID NO: 2272) I087B02 26-35 50-66  99-113 PFYDTLTSYVFRPDL(SEQ ID NO: 2281) I087B03 26-35 50-66  99-113 PFYDTLTSYVKSMPT(SEQ ID NO: 2305) I087B04 26-35 50-66  99-113 PFYDTLTSYVPFLYC(SEQ ID NO: 2292) I087B05 26-35 50-66  99-113 PFYDTLTSYVPVPST(SEQ ID NO: 2270) I087B06 26-35 50-66  99-113 PFYDTLTSYVGIHGL(SEQ ID NO: 2282) I087B08 26-35 50-66  99-114 PFYDTLTSYVPCSPPR(SEQ ID NO: 2261) I087B09 26-35 50-66  99-113 PFYDTLTSYVCYPPA(SEQ ID NO: 2240) I087C02 26-35 50-66  99-113 PFYDTLTSYVLPLLS(SEQ ID NO: 2224) I087C05 26-35 50-66  99-113 PFYDTLTSYVALYRL(SEQ ID NO: 2234) I087C06 26-35 50-66  99-113 PFYDTLTSYVRASFS(SEQ ID NO: 2271) I087C07 26-35 50-66  99-113 PFYDTLTSYVCTPVP(SEQ ID NO: 2319) I087C08 26-35 50-66  99-114 PFYDTLTSYVWPSFFS(SEQ ID NO: 2277) I087D01 26-35 50-66  99-113 PFYDTLTSYVTPRGY(SEQ ID NO: 2275) I087B02 26-35 50-66  99-113 PFYDTLTSYVSSLLS(SEQ ID NO: 2213) I087D03 26-35 50-66  99-114 PFYDTLTSYVPLLPLC(SEQ ID NO: 2263) I087D05 26-35 50-66  99-114 PFYDTLTSYVPPPSFL(SEQ ID NO: 2266) I087D07 26-35 50-66  99-113 PFYDTLTSYVPTSTT(SEQ ID NO: 2269) I087D09 26-35 50-66  99-114 PFYDTLTSYVISCSWA(SEQ ID NO: 2299) I087E04 26-35 50-66  99-114 PFYDTLTSYVSALPPP(SEQ ID NO: 2274) I087E05 26-35 50-66  99-113 PFYDTLTSYVCRHLF(SEQ ID NO: 2236) I087E10 26-35 50-66  99-114 PFYDTLTSYVVSFPSL(SEQ ID NO: 2307) I087F02 26-35 50-66  99-114 PFYDTLTSYVMGVTPS(SEQ ID NO: 2322) I087F04 26-35 50-66  99-114 PFYDTLTSYVLFRPVL(SEQ ID NO: 2326) I087F05 26-35 50-66  99-113 PFYDTLTSYVPSVGG(SEQ ID NO: 2267) I087F07 26-35 50-66  99-113 PFYDTLTSYVPPTRH(SEQ ID NO: 2286) I087F08 26-35 50-66  99-113 PFYDTLTSYVLRSRD(SEQ ID NO: 2243) I087F09 26-35 50-66  99-113 PFYDTLTSYVPLLPP(SEQ ID NO: 2310) I087G05 26-35 50-66  99-113 PFYDTLTSYVLRCVL(SEQ ID NO: 2239) I087G06 26-35 50-66  99-113 PFYDTLTSYVHPSRS(SEQ ID NO: 2285) I087G07 26-35 50-66  99-114 PFYDTLTSYVLRLPPQ(SEQ ID NO: 2241) I087G09 26-35 50-66  99-113 PFYDTLTSYVGPYGT(SEQ ID NO: 2284) I087G10 26-35 50-66  99-113 PFYDTLTSYVTTPCT(SEQ ID NO: 2276) I087H02 26-35 50-66  99-110 ASYLSTSSSLDN(SEQ ID NO: 2265) I088A01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088A03 26-35 50-66  99-114 PFYDTLTSYVIPFLPL(SEQ ID NO: 2290) I088A04 26-35 50-66  99-114 PFYDTLTSYVLHIYPH(SEQ ID NO: 2335) I088A08 26-35 50-66  99-114 PFYDTLTNYVFEYYAS(SEQ ID NO: 2323) I088A09 26-35 50-66  99-114 PFYDTLTSYVILYYLH(SEQ ID NO: 2295) I088A10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088A11 26-35 50-66  99-114 PFYDTLTSYVLMYFPH(SEQ ID NO: 2220) I088A12 26-35 50-66  99-114 PFYDTLTSYVLFFYPL(SEQ ID NO: 2325) I088B01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088B02 26-35 50-66  99-114 PFYDTLTSYVFDYYAS(SEQ ID NO: 2244) I088B03 26-35 50-66  99-114 PFYDTLTSYVIPFLPL(SEQ ID NO: 2290) I088B05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088B06 26-35 50-66  99-114 PFYDTLTSYVFEYYSL(SEQ ID NO: 2324) I088B07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088B08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088B09 26-35 50-66  99-114 PFYDTLTSYVLEFYLL(SEQ ID NO: 2303) I088B10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088B12 26-35 50-66  99-113 PFYDTLTSYVLPLDS(SEQ ID NO: 2223) I088C01 26-35 50-66  99-114 PFYDTLTSYVLYFYPS(SEQ ID NO: 2317) I088C03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088C09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088C12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088D01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088D03 26-35 50-66  99-114 PFYDTLTSYVLHYYAL(SEQ ID NO: 2215) I088D04 26-35 50-66  99-113 PFYDTLTSYVLPPSV(SEQ ID NO: 2225) I088D07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088D08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088D11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088E01 23-32 47-63  96-111 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088E02 26-35 50-66  99-114 PFYDTLTSYVLHYYLY(SEQ ID NO: 2216) I088E03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088E04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088E08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088E10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088E11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088F07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088G02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088G03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088G07 26-35 50-66  99-114 PFYDTLTSYVFHYYPL(SEQ ID NO: 2260) I088G09 26-35 50-66  99-114 PFYDTLTSYVFPVYYL(SEQ ID NO: 2264) I088G10 26-35 50-66  99-114 PFYDTLTSYVLHFIDH(SEQ ID NO: 2301) I088H05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I088H07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092A03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092A05 26-35 50-66  99-114 PFYDTLTSYVFHYYDV(SEQ ID NO: 2258) I092A06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092A08 26-35 50-66  99-114 PFYDTLTSYVHEFFSL(SEQ ID NO: 2283) I092A10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092A11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092B01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092B02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092B04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092B05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092B10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092B12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092C01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092C02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092C07 26-35 50-66  99-113 PFYDTLTSYVLALDL(SEQ ID NO: 2328) I092C08 26-35 50-66  99-114 PFYDTLTSYVFGYYSL(SEQ ID NO: 2254) I092C12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092D01 26-35 50-66  99-114 PFYDTLTSYVLKYYTD(SEQ ID NO: 2226) I092D07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092D09 26-35 50-66  99-114 PFYDTLTSYVMHAYPL(SEQ ID NO: 2255) I092D10 26-35 50-66  99-114 PFYDTLTSYVFHYLPV(SEQ ID NO: 2256) I092D11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092E01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092E03 26-35 50-66  99-114 PFYDTLTSYAFQYFDH(SEQ ID NO: 2230) I092E04 26-35 50-66  99-114 PFYDTLTSYVFEYFSV(SEQ ID NO: 2248) I092E07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092E10 26-35 50-66  99-114 PFYDTLTSYVLFYYPL(SEQ ID NO: 2327) I092E11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092F01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092F02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092F05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092F07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092F08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092F11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092F12 26-35 50-66  99-114 PFYDTLTSYVLAYYPD(SEQ ID NO: 2306) I092G01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092G05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092G10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I092H01 26-35 50-66  99-110 ASYLSTSSSLDN(SEQ ID NO: 2265) I093A06 26-35 50-66  99-114 PFYDTLTSYVLPVYDH(SEQ ID NO: 2334) I093A09 26-35 50-66  99-114 PFYDTLTSYVFQYFAH(SEQ ID NO: 2268) I093A11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093Al2 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093B02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093B05 26-35 50-66  99-114 PFYDTLTSYVIFYYPT(SEQ ID NO: 2289) I093B06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093B09 26-35 50-66  99-114 PFYDTLTSYVLEVYHP(SEQ ID NO: 2318) I093B12 26-35 50-66  99-114 PFYDTLTSYVFAPLVT(SEQ ID NO: 2242) I093C02 26-35 50-66  99-114 PFYDTLTSYVLHAYAF(SEQ ID NO: 2332) I093C03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093C05 26-35 50-66  99-114 PFYDTLTSYVILYYLH(SEQ ID NO: 2295) I093D05 26-35 50-66  99-114 PFYDTLTSYVFEFLPL(SEQ ID NO: 2245) I093D08 26-35 50-66  99-114 PFYDTLTSYVRPFYAH(SEQ ID NO: 2273) I093D10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093D12 26-35 50-66  99-114 PFYDTLTSYVLHFYRV(SEQ ID NO: 2302) I093E01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093E02 26-35 50-66  99-114 PFYDTLTSYVIQYFDH(SEQ ID NO: 2297) I093E05 26-35 50-66  99-114 PFYDTLTSYVHEFFSL(SEQ ID NO: 2283) I093E08 26-35 50-66  99-114 PFYDTLTSYVMQFFPT(SEQ ID NO: 2321) I093E10 26-35 50-66  99-114 PFYDTLTSYVLSFYPV(SEQ ID NO: 2246) I093F01 26-35 50-66  99-114 PFDYTLTSYVLYYYAF(SEQ ID NO: 2251) I093F03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093F05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093F08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093F11 26-35 50-66  99-114 PFYDTLTSYVLHFYPL(SEQ ID NO: 2333) I093G07 26-35 50-66  99-114 PFYDTLTSYVLQYYVL(SEQ ID NO: 2237) I093G11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093G12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I093H06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094A08 26-35 50-66  99-114 PFYDTLTSYVFQYFDY(SEQ ID NO: 2280) I094B07 26-35 50-66  99-114 PFYDTLTSYVLPVWVS(SEQ ID NO: 2228) I094B08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094B12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094C11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094C12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094D06 26-35 50-66  99-114 PFYDTLTSYVIEYYPV(SEQ ID NO: 2288) I094D07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094D08 26-35 50-66  99-114 PFYDTLTSYVLHYLPL(SEQ ID NO: 2314) I094D09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094D10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094D11 26-35 50-66  99-114 PFYDTLTSYVFHFYPV(SEQ ID NO: 2218) I094E04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094E08 26-35 50-66  99-114 PFYDTLTSYVLEAFSL(SEQ ID NO: 2311) I094F04 26-35 50-66  99-114 PFYDTLTSYVFGFYPF(SEQ ID NO: 2252) I094F05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094F10 26-35 50-66  99-114 PIYDTLTSYVFQYFDH(SEQ ID NO: 2278) I094F11 26-35 50-66  99-114 PFYDTLTSYVLWYYQD(SEQ ID NO: 2249) I094F12 26-35 50-66  99-114 PFYDTLTSYVIPFYPL(SEQ ID NO: 2296) I094G06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I094G10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095A04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095Al2 26-35 50-66  99-114 PFYDTLTSYVLEYFPL(SEQ ID NO: 2320) I095B04 26-35 50-66  99-114 PFYDTLTSYVLEFFPA(SEQ ID NO: 2312) I095B09 26-35 50-66  99-114 PFYDTLTSYVIEYLPL(SEQ ID NO: 2287) I095B10 26-35 50-66  99-114 PFYDTLTSYVLHYYSA(SEQ ID NO: 2217) I095C02 26-35 50-66  99-114 PFYDTLTSYVLFYYTA(SEQ ID NO: 2331) I095C05 26-35 50-66  99-114 PFYDTLTSYVLHYLPV(SEQ ID NO: 2337) I095C07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095C08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095C09 26-35 50-66  99-114 PFYDTLTSYVMHYYPT(SEQ ID NO: 2259) I095D01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095D02 26-35 50-66  99-114 PFYDTLTSYVLQYFRY(SEQ ID NO: 2235) I095D03 26-35 50-66  99-114 PFYDTLTSYVLQVFDT(SEQ ID NO: 2233) I095D05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095D09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095E01 26-35 50-66  99-114 PFYDTLTSYVLDYYSS(SEQ ID NO: 2309) I095E05 26-35 50-66  99-114 PFYDALTSYVFQYFDH(SEQ ID NO: 2221) I095E12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095F06 26-35 50-66  99-114 PFYDTLTSYVFPFYPH(SEQ ID NO: 2262) I095F09 26-35 50-66  99-114 PFYDTLTSYVIGFYPV(SEQ ID NO: 2291) I095G06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I095G09 26-35 50-66  99-114 PFYDTLTSYVMDFYSV(SEQ ID NO: 2253) I095G11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096A01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096A10 26-35 50-66  99-114 PFYDTLTSYVLPFYAL(SEQ ID NO: 2222) I096B01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096B03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096C01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096C06 26-35 50-66  99-114 PFYDTLTSYVLPYLTH(SEQ ID NO: 2229) I096C09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096D01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096D02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096D05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096D06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096D09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096E02 26-35 50-66  99-114 PFYDTLTSYVLGFYPV(SEQ ID NO: 2329) I096E06 26-35 50-66  99-114 PFYDTLTSYVLHYHTH(SEQ ID NO: 2336) I096E11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096F02 26-35 50-66  99-114 PFYDTLTSYVIHFLPL(SEQ ID NO: 2330) I096G01 26-35 50-66  99-114 PFYDTLTSYVIPFLPL(SEQ ID NO: 2290) I096G02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096G05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096G07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I096G09 26-35 50-66  99-114 PFYDTLTSYVMHYLPV(SEQ ID NO: 2257) I096G12 26-35 50-66  99-114 PFYDTLTSYVLEFFSH(SEQ ID NO: 2315) I096H01 26-35 50-66  99-110 ASYLSTSSSLDN(SEQ ID NO: 2265) I097A04 26-35 50-66  99-114 PFYDTLTSYVIHYLVT(SEQ ID NO: 2294) I097A06 26-35 50-66  99-114 PFYDTLTSYVLPYYTL(SEQ ID NO: 2231) I097A09 26-35 50-66  99-114 PFYDTLTSYVLHYYPI(SEQ ID NO: 2298) I097B02 26-35 50-66  99-114 PFYDTLTSYVLWFYPL(SEQ ID NO: 2247) I097B09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097B10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097B11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097C05 26-35 50-66  99-114 PFYDTLTSYVLHYYTH(SEQ ID NO: 2219) I097C09 26-35 50-66  99-114 PFYDTLTSYVLHYYAY(SEQ ID NO: 2316) I097C11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097D05 26-35 50-66  99-114 PFYDTLTSYVIHFYSL(SEQ ID NO: 2293) I097D06 26-35 50-66  99-114 PFYDTLTSYVFGFFPH(SEQ ID NO: 2300) I097E01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097E04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097E08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097E09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097F09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I097G10 26-35 50-66  99-114 PFYDTLTSYVFQYFAH(SEQ ID NO: 2268) I097H02 26-35 50-66  99-110 ASYLSTSSSLDN(SEQ ID NO: 2265) I098A04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I098A05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I098B08 26-35 50-66  99-114 PFYDTLTSYVLDFYSV(SEQ ID NO: 2308) I098C01 26-35 50-66  99-114 PIYDTLTSYVFQYFDH(SEQ ID NO: 2278) I098C04 26-35 50-66  99-114 PFYDTLTSYVLYYYAF(SEQ ID NO: 2251) I098F11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I098F12 26-35 50-66  99-114 PFYDTLTSYVFFFYPF(SEQ ID NO: 2250) I098G02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I098G12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I098H05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I101A01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I101B04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I101B06 26-35 50-66  99-114 PFYDTLTSYVIPFLTH(SEQ ID NO: 2304) I101D04 26-35 50-66  99-114 PFYDTLTSYVLEFFPD(SEQ ID NO: 2313) I101D07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I101E09 26-35 50-66  99-114 PFYDTLTSYVFQYFDR(SEQ ID NO: 2279) I101E12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I101G02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I101G11 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I102C03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I102E09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I102F02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I102G08 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I102G09 26-35 50-66  99-114 PFYDTLTSYVLHYYAH(SEQ ID NO: 2214) I106A09 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I106B02 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I106B06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I106C07 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I106E05 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I106E12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I106G01 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I106G03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I109B06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I109D12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I109E12 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I109G06 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I109H04 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I110B03 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I112D09 26-35 50-66  99-114 PFYDTLTSYGFQYFDH(SEQ ID NO: 2232) I112F10 26-35 50-66  99-114 PFYDTLTSYVFQYFDH(SEQ ID NO: 2137) I089F12 26-35 50-66  99-112 SRDLLLFPHHGLDS(SEQ ID NO: 2146) I105E12 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I108D08 26-35 50-66  99-112 SRDLLLFPLAPLYP(SEQ ID NO: 2148) I108E06 26-35 50-66  99-112 SRDLLLFPHHGLDV(SEQ ID NO: 2151) I113E07 26-35 50-66  99-112 SRDLLLFPHHSLDL(SEQ ID NO: 2152) I114G05 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I116A01 26-35 50-66  99-112 SRDLLLFPHHALSP(SEQ ID NO: 2149) I116A09 26-35 50-66  99-112 SRYLLLFPHHSFDL(SEQ ID NO: 2150) I116C11 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I085A01 26-35 50-66  99-112 SRDLLLFPHDHLLF(SEQ ID NO: 2602) I085A02 26-35 50-66  99-112 SRDLLLFPSDPLGF(SEQ ID NO: 2639) I085A03 26-35 50-66  99-112 SRDLLLFPTHPLSF(SEQ ID NO: 2561) I085A04 26-35 50-66  99-112 SRDLLLFPLAPLFF(SEQ ID NO: 2550) I085A05 26-35 50-66  99-112 SRDLLLFPSDPLSL(SEQ ID NO: 2659) I085A06 26-35 50-66  99-112 SRDLLLFPSAPLSF(SEQ ID NO: 2611) I085A07 26-35 50-66  99-112 SRDLLLFPASPLSF(SEQ ID NO: 2390) I085A09 26-35 50-66  99-111 SRDLLLFPNDALS(SEQ ID NO: 2632) I085A10 26-35 50-66  99-112 SRDLLLFPSAPLRF(SEQ ID NO: 2609) I085A11 26-35 50-66  99-111 SRDLLLFPHDPLE(SEQ ID NO: 2363) I085B01 26-35 50-66  99-112 SRDLLLFPQSPLYP(SEQ ID NO: 2466) I085B02 26-35 50-66  99-112 SRDLLLFPHSSLVF(SEQ ID NO: 2392) I085B03 26-35 50-66  99-112 SRDLLLFPYDPLLF(SEQ ID NO: 2638) I085B04 26-35 50-66  99-112 SRDLLLFPHAPYLF(SEQ ID NO: 2589) I085B05 26-35 50-66  99-112 SRDLLLFPHAPLSP(SEQ ID NO: 2573) I085B06 26-35 50-66  99-112 SRDLLLFPLSPLSF(SEQ ID NO: 2574) I085B07 26-35 50-66  99-112 SRDLLLFPDFPMAP(SEQ ID NO: 2433) I085B10 26-35 50-66  99-111 SRDLLLFPHSPLY(SEQ ID NO: 2470) I085B12 26-35 50-66  99-112 SRDLLLFPQDPLSP(SEQ ID NO: 2372) I085C02 26-35 50-66  99-112 SRDLLLFPDDPLLS(SEQ ID NO: 2430) I085C03 26-35 50-66  99-112 SRDLLLFPHGPLLI(SEQ ID NO: 2400) I085C05 26-35 50-66  99-112 SRDLLLFPGSPLLF(SEQ ID NO: 2491) I085C06 26-35 50-66  99-112 SRDLLLFPTAALSF(SEQ ID NO: 2341) I085C07 26-35 50-66  99-112 SRDLLLFPHTPLRF(SEQ ID NO: 2375) I085C09 26-35 50-66  99-111 SRDLLLFPHSPLT(SEQ ID NO: 2468) I085C10 26-35 50-66  99-112 SRDLLLFPFSPLLF(SEQ ID NO: 2471) I085C12 26-35 50-66  99-112 SRDLLLFPSHPLFF(SEQ ID NO: 2680) I085D01 26-35 50-66  99-112 SRDLLLFPRPLLLF(SEQ ID NO: 2548) I085D02 26-35 50-66  99-112 SRDLLLFPSQYLDF(SEQ ID NO: 2523) I085D03 26-35 50-66  99-112 SRDLLLFPSSPLLF(SEQ ID NO: 2713) I085D04 26-35 50-66  99-112 SRDLLLFPYFPLVF(SEQ ID NO: 2646) I085D06 26-35 50-66  99-112 SRDLLLFPGSPLLD(SEQ ID NO: 2488) I085D07 26-35 50-66  99-111 SRDLLLFPQAPLLF(SEQ ID NO: 2694) I085D08 26-35 50-66  99-112 SRDLLLFPSHYLSP(SEQ ID NO: 2477) I085D09 26-35 50-66  99-112 SRDLLLFPQTPLFP(SEQ ID NO: 2467) I085D10 26-35 50-66  99-112 SRDLLLFPHSPLHP(SEQ ID NO: 2563) I085D11 26-35 50-66  99-112 SRDLLLFPHAPLAP(SEQ ID NO: 2510) I085D12 26-35 50-66  99-112 SRDLLLFPHTTLRF(SEQ ID NO: 2495) I085E01 26-35 50-66  99-112 SRDLLLFPYAVLHF(SEQ ID NO: 2620) I085E02 26-35 50-66  99-112 SRDLLLFPTSPLRL(SEQ ID NO: 2575) I085E07 26-35 50-66  99-112 SRDLLLFPSDALSF(SEQ ID NO: 2568) I085E08 26-35 50-66  99-112 SRDLLLFPNAPLDP(SEQ ID NO: 2603) I085E09 26-35 50-66  99-112 SRDLLLFPHDPPRF(SEQ ID NO: 2628) I085E10 26-35 50-66  99-112 SRDLLLFPSEPLWP(SEQ ID NO: 2668) I085E11 26-35 50-66  99-112 SRDLLLFPSSPLSN(SEQ ID NO: 2716) I085E12 26-35 50-66  99-112 SRDLLLFPHLPLTP(SEQ ID NO: 2431) I085F01 26-35 50-66  99-111 SRDLLLFPRSPLLF(SEQ ID NO: 2551) I085F02 26-35 50-66  99-112 SRDLLLFPTSPLQL(SEQ ID NO: 2376) I085F03 26-35 50-66  99-112 SRDLLLFPYTPLLF(SEQ ID NO: 2682) I085F04 26-35 50-66  99-112 SRDLLLFPSSPLAF(SEQ ID NO: 2707) I085F05 26-35 50-66  99-112 SRDLLLFPHDPLYF(SEQ ID NO: 2706) I085F06 26-35 50-66  99-112 SRDLLLFPSAHLLF(SEQ ID NO: 2586) I085F07 26-35 50-66  99-112 SRDLLLFPAGPLRF(SEQ ID NO: 2410) I085F09 26-35 50-66  99-112 SRDLLLFPDHAFFV(SEQ ID NO: 2439) I085F10 26-35 50-66  99-112 SRDLLLFPSDSGFA(SEQ ID NO: 2662) I085F11 26-35 50-66  99-112 SRDLLLFPSSYLEF(SEQ ID NO: 2339) I085F12 26-35 50-66  99-111 SRDLLLFPRDPLII(SEQ ID NO: 2558) I085G01 26-35 50-66  99-112 SRDLLLFPSAPLHP(SEQ ID NO: 2605) I085G02 26-35 50-66  99-112 SRDLLLFPNAPLLL(SEQ ID NO: 2613) I085G03 26-35 50-66  99-112 SRDLLLFPAAPLLF(SEQ ID NO: 2403) I085G04 26-35 50-66  99-112 SRDLLLFPSAPLDP(SEQ ID NO: 2601) I085G07 26-35 50-66  99-112 SRDLLLFPNAVLDI(SEQ ID NO: 2629) I085G08 26-35 50-66  99-112 SRDLLLFPSEPLFF(SEQ ID NO: 2664) I085G09 26-35 50-66  99-112 SRDLLLFPSSVLWP(SEQ ID NO: 2338) I085G10 26-35 50-66  99-111 SRDLLLFPHAPLQ(SEQ ID NO: 2554) I085G11 26-35 50-66  99-112 SRDLLLFPDSPLAP(SEQ ID NO: 2445) I085G12 26-35 50-66  99-112 SRDLLLFPSSPLHP(SEQ ID NO: 2576) I085H10 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I086A03 26-35 50-66  99-112 SRDLLLFPFSMPLTF(SEQ ID NO: 2695) I086A04 26-35 50-66  99-112 SRDLLLFPHSILHP(SEQ ID NO: 2438) I086A05 26-35 50-66  99-111 SRDLLLFPHAPLSH(SEQ ID NO: 2569) I086A07 26-35 50-66  99-112 SRDLLLFPDAALRF(SEQ ID NO: 2421) I086A09 26-35 50-66  99-112 SRDLLLFPSSHLSF(SEQ ID NO: 2704) I086A10 26-35 50-66  99-112 SRDLLLFPSAPLSS(SEQ ID NO: 2624) I086A11 26-35 50-66  99-112 SRDLLLFPHAPLTP(SEQ ID NO: 2577) I086Al2 26-35 50-66  99-112 SRDLLLFPYDPLHS(SEQ ID NO: 2635) I086B02 26-35 50-66  99-112 SRDLLLFPHFPLHP(SEQ ID NO: 2348) I086B03 26-35 50-66  99-112 SRDLLLFPAHPLLF(SEQ ID NO: 2412) I086B05 26-35 50-66  99-112 SRDLLLFPFEPLII(SEQ ID NO: 2457) I086B06 26-35 50-66  99-112 SRDLLLFPASPLNP(SEQ ID NO: 2364) I086B07 26-35 50-66  99-112 SRDLLLFPSSPLYF(SEQ ID NO: 2720) I086B09 26-35 50-66  99-112 SRDLLLFPTSPLSF(SEQ ID NO: 2579) I086B10 26-35 50-66  99-112 SRDLLLFPDDGLSS(SEQ ID NO: 2428) I086B11 26-35 50-66  99-112 SRDLLLFPISPLCF(SEQ ID NO: 2530) I086C03 26-35 50-66  99-112 SRDLLLFPTAPLYG(SEQ ID NO: 2535) I086C05 26-35 50-66  99-112 SRDLLLFPHHSLFF(SEQ ID NO: 2427) I086C07 26-35 50-66  99-112 SRDLLLFPQGPLRF(SEQ ID NO: 2440) I086C08 26-35 50-66  99-112 SRDLLLFPAAPLAF(SEQ ID NO: 2401) I086C09 26-35 50-66  99-112 SRDLLLFPHLPLLF(SEQ ID NO: 2350) I086C10 26-35 50-66  99-112 SRDLLLFPTFPLIF(SEQ ID NO: 2541) I086C11 26-35 50-66  99-112 SRDLLLFPDDPLSF(SEQ ID NO: 2432) I086C12 26-35 50-66  99-112 SRDLLLFPTDSLLF(SEQ ID NO: 2622) I086D01 26-35 50-66  99-112 SRDLLLFPSAPLTP(SEQ ID NO: 2630) I086D04 26-35 50-66  99-112 SRDLLLFPYAPLYD(SEQ ID NO: 2697) I086D05 26-35 50-66  99-112 SRDLLLFPHSPLSF(SEQ ID NO: 2461) I086D06 26-35 50-66  99-112 SRDLLLFPTAPLDL(SEQ ID NO: 2379) I086D07 26-35 50-66  99-112 SRDLLLFPHTHLTF(SEQ ID NO: 2365) I086D08 26-35 50-66  99-112 SRDLLLFPHSSLDF(SEQ ID NO: 2473) I086D09 26-35 50-66  99-112 SRDLLLFPNHPMFP(SEQ ID NO: 2665) I086D10 26-35 50-66  99-112 SRDLLLFPLSSLEF(SEQ ID NO: 2587) I086D11 26-35 50-66  99-112 SRDLLLFPNAPLHP(SEQ ID NO: 2610) I086D12 26-35 50-66  99-112 SRDLLLFPRAHLRF(SEQ ID NO: 2469) I086E02 26-35 50-66  99-112 SRDLLLFPYDPLHF(SEQ ID NO: 2621) I086E03 26-35 50-66  99-112 SRDLLLFPHDALQS(SEQ ID NO: 2598) I086E05 26-35 50-66  99-112 SRDLLLFPRTPLTF(SEQ ID NO: 2567) I086E07 26-35 50-66  99-112 SRDLLLFPAAHLSF(SEQ ID NO: 2398) I086E08 26-35 50-66  99-112 SRDLLLFPRAPLLF(SEQ ID NO: 2490) I086E09 26-35 50-66  99-112 SRDLLLFPFSPLAP(SEQ ID NO: 2464) I086E10 26-35 50-66  99-112 SRDLLLFPRAPLDF(SEQ ID NO: 2367) I086E12 26-35 50-66  99-112 SRDLLLFPTAPLRF(SEQ ID NO: 2522) I086F02 26-35 50-66  99-112 SRDLLLFPSSPLRI(SEQ ID NO: 2714) I086F05 26-35 50-66  99-112 SRDLLLFPTEPLQF(SEQ ID NO: 2540) I086F08 26-35 50-66  99-112 SRDLLLFPSDPLSA(SEQ ID NO: 2643) I086F09 26-35 50-66  99-112 SRDLLLFPYNPPIF(SEQ ID NO: 2653) I086F11 26-35 50-66  99-112 SRDLLLFPHTPLLF(SEQ ID NO: 2489) I086G03 26-35 50-66  99-112 SRDLLLFPHAPLDL(SEQ ID NO: 2513) I086G04 26-35 50-66  99-112 SRDLLLFPFDPLLI(SEQ ID NO: 2454) I086G05 26-35 50-66  99-112 SRDLLLFPTDALRI(SEQ ID NO: 2537) I086G06 26-35 50-66  99-112 SRDLLLFPAAPLTP(SEQ ID NO: 2407) I086G07 26-35 50-66  99-112 SRDLLLFPEGPLLF(SEQ ID NO: 2448) I086G09 26-35 50-66  99-112 SRDLLLFPYAPLSF(SEQ ID NO: 2385) I086G10 26-35 50-66  99-112 SRDLLLFPADSLSF(SEQ ID NO: 2391) I086H05 26-35 50-66  99-112 SRDLLLFPYSPLTH(SEQ ID NO: 2679) I089A01 26-35 50-66  99-111 SRDLLLFPHDPLI(SEQ ID NO: 2612) I089A03 26-35 50-66  99-112 SRDLLLFPLTPLLI(SEQ ID NO: 2590) I089A06 26-35 50-66  99-112 SRDLLLFPHTPLHF(SEQ ID NO: 2485) I089A07 26-35 50-66  99-112 SRDLLLFPTDALYF(SEQ ID NO: 2539) I089A08 26-35 50-66  99-112 SRDLLLFPYTPLLF(SEQ ID NO: 2682) I089A10 26-35 50-66  99-112 SRDLLLFPHQPLTF(SEQ ID NO: 2436) I089A11 26-35 50-66  99-112 SRDLLLFPRTYLDF(SEQ ID NO: 2572) I089B01 26-35 50-66  99-112 SRDLLLFPHSPLHS(SEQ ID NO: 2450) I089B02 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I089B03 26-35 50-66  99-112 SRDLLLFPTSPLQP(SEQ ID NO: 2528) I089B04 26-35 50-66  99-112 SRDLLLFPTHPLLF(SEQ ID NO: 2556) I089B05 26-35 50-66  99-112 SRDLLLFPSSPLIF(SEQ ID NO: 2712) I089B06 26-35 50-66  99-112 SRDLLLFPMAPLSP(SEQ ID NO: 2596) I089B07 26-35 50-66  99-112 SRDLLLFPYSGLDA(SEQ ID NO: 2374) I089B08 26-35 50-66  99-112 SRDLLLFPAAPLSP(SEQ ID NO: 2405) I089B09 26-35 50-66  99-112 SRDLLLFPKSPILF(SEQ ID NO: 2384) I089B10 26-35 50-66  99-112 SRDLLLFPTSPLFF(SEQ ID NO: 2571) I089B11 26-35 50-66  99-112 SRDLLLFPNSPLFP(SEQ ID NO: 2388) I089C01 26-35 50-66  99-112 SRDLLLFPHYFMDV(SEQ ID NO: 2133) I089C02 26-35 50-66  99-112 SRDLLLFPRSPLLF(SEQ ID NO: 2551) I089C03 26-35 50-66  99-112 SRDLLLFPYHPLLF(SEQ ID NO: 2532) I089C05 26-35 50-66  99-112 SRDLLLFPSSALRF(SEQ ID NO: 2722) I089C06 26-35 50-66  99-112 SRDLLLFPSPYLSF(SEQ ID NO: 2701) I089C07 26-35 50-66  99-112 SRDLLLFPQAPLFD(SEQ ID NO: 2683) I089C09 26-35 50-66  99-112 SRDLLLFPHAPFTF(SEQ ID NO: 2507) I089D01 26-35 50-66  99-112 SRDLLLFPHAPLVL(SEQ ID NO: 2581) I089D02 26-35 50-66  99-112 SRDLLLFPHYFMDV(SEQ ID NO: 2133) I089D03 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I089D04 26-35 50-66  99-112 SRDLLLFPSSPLSP(SEQ ID NO: 2717) I089D05 26-35 50-66  99-112 SRDLLLFPHAPLFT(SEQ ID NO: 2546) I089D07 26-35 50-66  99-112 SRDLLLFPNDPLLI(SEQ ID NO: 2634) I089D08 26-35 50-66  99-111 SRDLLLFPHAPLQ(SEQ ID NO: 2554) I089D09 26-35 50-66  99-112 SRDLLLFPSHAFHE(SEQ ID NO: 2677) I089D11 26-35 50-66  99-112 SRDLLLFPNHPLYP(SEQ ID NO: 2663) I089E01 26-35 50-66  99-112 SRDLLLFPYSPLFP(SEQ ID NO: 2657) I089E02 26-35 50-66  99-112 SRDLLLFPQDPLHP(SEQ ID NO: 2346) I089E03 26-35 50-66  99-112 SRDLLLFPDAPLFP(SEQ ID NO: 2423) I089E04 26-35 50-66  99-112 SRDLLLFPHSPLLI(SEQ ID NO: 2453) I089E06 26-35 50-66  99-112 SRDLLLFPGSPLLF(SEQ ID NO: 2491) I089E09 26-35 50-66  99-112 SRDLLLFPSSPLTF(SEQ ID NO: 2718) I089E10 26-35 50-66  99-112 SRDLLLFPTQPLSF(SEQ ID NO: 2566) I089E11 26-35 50-66  99-112 SRDLLLFPLSPLWP(SEQ ID NO: 2578) I089F01 26-35 50-66  99-112 SRDLLLFPTFPLLF(SEQ ID NO: 2380) I089F03 26-35 50-66  99-112 SRDLLLFPHDPLLL(SEQ ID NO: 2580) I089F04 26-35 50-66  99-112 SRDLLLFPYSPLLF(SEQ ID NO: 2670) I089F05 26-35 50-66  99-112 SRDLLLFPHSPLRI(SEQ ID NO: 2459) I089F06 26-35 50-66  99-112 SRDLLLFPRAPLLF(SEQ ID NO: 2490) I089F08 26-35 50-66  99-112 SRDLLLFPRTPLTF(SEQ ID NO: 2567) I089F09 26-35 50-66  99-112 SRDLLLFPLAPLSF(SEQ ID NO: 2555) I089F10 26-35 50-66  99-112 SRDLLLFPNQPLSF(SEQ ID NO: 2667) I089F11 26-35 50-66  99-112 SRDLLLFPLEPMHF(SEQ ID NO: 2565) I089G01 26-35 50-66  99-112 SRDLLLFPSAPLTF(SEQ ID NO: 2626) I089G02 26-35 50-66  99-112 SRDLLLFPSHPLLF(SEQ ID NO: 2687) I089G03 26-35 50-66  99-112 SRDLLLFPRTPLVF(SEQ ID NO: 2721) I089G05 26-35 50-66  99-112 SRDLLLFPGSPLTF(SEQ ID NO: 2389) I089G06 26-35 50-66  99-112 SRDLLLFPTAPLLF(SEQ ID NO: 2514) I089G07 26-35 50-66  99-112 SRDLLLFPSAPLDF(SEQ ID NO: 2597) I089G08 26-35 50-66  99-112 SRDLLLFPSHPLSF(SEQ ID NO: 2688) I089G11 26-35 50-66  99-112 SRDLLLFPSFPLLF(SEQ ID NO: 2671) I089H10 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I090A02 26-35 50-66  99-112 SRDLLLFPAKPLLF(SEQ ID NO: 2416) I090A03 26-35 50-66  99-112 SRDLLLFPNSTLSF(SEQ ID NO: 2678) I090A04 26-35 50-66  99-112 SRDLLLFPDAPLTF(SEQ ID NO: 2426) I090A05 26-35 50-66  99-112 SRDLLLFPHEPLLI(SEQ ID NO: 2648) I090A06 26-35 50-66  99-112 SRDLLLFPTYPLSF(SEQ ID NO: 2600) I090A07 26-35 50-66  99-112 SRDLLLFPTEPLVL(SEQ ID NO: 2479) I090A08 26-35 50-66  99-112 SRDLLLFPTYPLHF(SEQ ID NO: 2584) I090B01 26-35 50-66  99-112 SRDLLLFPHDPLTF(SEQ ID NO: 2627) I090B03 26-35 50-66  99-112 SRDLLLFPQAPLTN(SEQ ID NO: 2705) I090B04 26-35 50-66  99-112 SRDLLLFPHAPLEA(SEQ ID NO: 2520) I090B05 26-35 50-66  99-112 SRDLLLFPDHPLLF(SEQ ID NO: 2442) I090B06 26-35 50-66  99-112 SRDLLLFPRAPLSF(SEQ ID NO: 2496) I090B08 26-35 50-66  99-112 SRDLLLFPRGPLRF(SEQ ID NO: 2542) I090B11 26-35 50-66  99-112 SRDLLLFPFTPLTF(SEQ ID NO: 2474) I090B12 26-35 50-66  99-112 SRDLLLFPQHPLSP(SEQ ID NO: 2452) I090001 26-35 50-66  99-112 SRDLLLFPSAPIVF(SEQ ID NO: 2591) I090002 26-35 50-66  99-112 SRDLLLFPQAPLTF(SEQ ID NO: 2702) I090003 26-35 50-66  99-112 SRDLLLFPRAPLRF(SEQ ID NO: 2493) I090005 26-35 50-66  99-112 SRDLLLFPRTPLTF(SEQ ID NO: 2567) I090006 26-35 50-66  99-112 SRDLLLFPHAPLDF(SEQ ID NO: 2538) I090007 26-35 50-66  99-112 SRDLLLFPHAGFDS(SEQ ID NO: 2498) I090008 26-35 50-66  99-112 SRDLLLFPYSPLSF(SEQ ID NO: 2676) I090C10 26-35 50-66  99-112 SRDLLLFPGRPLTF(SEQ ID NO: 2358) I090D02 26-35 50-66  99-112 SRDLLLFPAEHLLF(SEQ ID NO: 2408) I090D03 26-35 50-66  99-112 SRDLLLFPTAPLHP(SEQ ID NO: 2351) I090D04 26-35 50-66  99-112 SRDLLLFPHEPLTA(SEQ ID NO: 2654) I090D05 26-35 50-66  99-112 SRDLLLFPHAPLFE(SEQ ID NO: 2529) I090D06 26-35 50-66  99-112 SRDLLLFPRAPLDF(SEQ ID NO: 2367) I090D07 26-35 50-66  99-112 SRDLLLFPFGTLRF(SEQ ID NO: 2462) I090D08 26-35 50-66  99-112 SRDLLLFPSSPLVF(SEQ ID NO: 2723) I090D09 26-35 50-66  99-112 SRDLLLFPRDPLAF(SEQ ID NO: 2505) I090D12 26-35 50-66  99-112 SRDLLLFPTSPLSF(SEQ ID NO: 2579) I090E04 26-35 50-66  99-112 SRDLLLFPHAPLLL(SEQ ID NO: 2552) I090E05 26-35 50-66  99-112 SRDLLLFPSAPISF(SEQ ID NO: 2588) I090E06 26-35 50-66  99-112 SRDLLLFPQGPLSF(SEQ ID NO: 2443) I090E07 26-35 50-66  99-112 SRDLLLFPGSPLHP(SEQ ID NO: 2484) I090E09 26-35 50-66  99-112 SRDLLLFPSDPLSF(SEQ ID NO: 2647) I090E11 26-35 50-66  99-112 SRDLLLFPHDGLAP(SEQ ID NO: 2700) I090E12 26-35 50-66  99-112 SRDLLLFPTSPLTF(SEQ ID NO: 2582) I090F01 26-35 50-66  99-112 SRDLLLFPNGPLHP(SEQ ID NO: 2649) I090F02 26-35 50-66  99-112 SRDLLLFPQAPLSF(SEQ ID NO: 2696) I090F03 26-35 50-66  99-112 SRDLLLFPTAPLSF(SEQ ID NO: 2526) I090F04 26-35 50-66  99-112 SRDLLLFPFFPLQF(SEQ ID NO: 2460) I090F05 26-35 50-66  99-112 SRDLLLFPLDPLHF(SEQ ID NO: 2359) I090F06 26-35 50-66  99-112 SRDLLLFPSEPLQL(SEQ ID NO: 2666) I090F07 26-35 50-66  99-112 SRDLLLFPFAPLRF(SEQ ID NO: 2451) I090F08 26-35 50-66  99-112 SRDLLLFPLHPLIF(SEQ ID NO: 2570) I090F09 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I090F10 26-35 50-66  99-112 SRDLLLFPRDPLRI(SEQ ID NO: 2527) I090F11 26-35 50-66  99-112 SRDLLLFPSNPLTF(SEQ ID NO: 2698) I090G01 26-35 50-66  99-112 SRDLLLFPTAPLEI(SEQ ID NO: 2347) I090G02 26-35 50-66  99-112 SRDLLLFPRDPLQF(SEQ ID NO: 2395) I090G04 26-35 50-66  99-112 SRDLLLFPHEPLAF(SEQ ID NO: 2633) I090G05 26-35 50-66  99-112 SRDLLLFPRAPLAF(SEQ ID NO: 2472) I090G06 26-35 50-66  99-112 SRDLLLFPYSPLAF(SEQ ID NO: 2656) I090G07 26-35 50-66  99-112 SRDLLLFPHTPLDS(SEQ ID NO: 2480) I090G08 26-35 50-66  99-112 SRDLLLFPHTPLTF(SEQ ID NO: 2492) I090G09 26-35 50-66  99-112 SRDLLLFPSEPLRI(SEQ ID NO: 2356) I090G10 26-35 50-66  99-112 SRDLLLFPTAPLDF(SEQ ID NO: 2343) I090G12 26-35 50-66  99-112 SRDLLLFPNRGLDL(SEQ ID NO: 2669) I091A02 26-35 50-66  99-112 SRDLLLFPYDPLFM(SEQ ID NO: 2724) I091A03 26-35 50-66  99-112 SRDLLLFPHAPLYP(SEQ ID NO: 2592) I091A06 26-35 50-66  99-112 SRDLLLFPSAPLAF(SEQ ID NO: 2594) I091A11 26-35 50-66  99-112 SRDLLLFPHSPITF(SEQ ID NO: 2441) I091B01 26-35 50-66  99-112 SRDLLLFPRYPLFF(SEQ ID NO: 2585) I091B02 26-35 50-66  99-112 SRDLLLFPYAPLDF(SEQ ID NO: 2361) I091B04 26-35 50-66  99-112 SRDLLLFPRDPLQF(SEQ ID NO: 2395) I091B05 26-35 50-66  99-112 SRDLLLFPHAPLEL(SEQ ID NO: 2475) I091B07 26-35 50-66  99-112 SRDLLLFPSAPLTF(SEQ ID NO: 2626) I091B10 26-35 50-66  99-112 SRDLLLFPTAPLAF(SEQ ID NO: 2342) I091B11 26-35 50-66  99-112 SRDLLLFPHSPLDF(SEQ ID NO: 2444) I091B12 26-35 50-66  99-112 SRDLLLFPSHPLTF(SEQ ID NO: 2690) I091C02 26-35 50-66  99-112 SRDLLLFPAHPLVI(SEQ ID NO: 2414) I091C03 26-35 50-66  99-112 SRDLLLFPQAPLYP(SEQ ID NO: 2378) I091C04 26-35 50-66  99-112 SRDLLLFPTAPLTF(SEQ ID NO: 2531) I091C05 26-35 50-66  99-112 SRDLLLFPTTPLHF(SEQ ID NO: 2583) I091C06 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I091C09 26-35 50-66  99-112 SRDLLLFPHHPLSF(SEQ ID NO: 2415) I091C11 26-35 50-66  99-112 SRDLLLFPYHSYDI(SEQ ID NO: 2650) I091C12 26-35 50-66  99-112 SRDLLLFPYATLSF(SEQ ID NO: 2618) I091D01 26-35 50-66  99-112 SRDLLLFPNSPLAP(SEQ ID NO: 2672) I091D02 26-35 50-66  99-112 SRDLLLFPYSPLQP(SEQ ID NO: 2673) I091D04 26-35 50-66  99-112 SRDLLLFPQGPLSF(SEQ ID NO: 2443) I091D05 26-35 50-66  99-112 SRDLLLFPHDPLAP(SEQ ID NO: 2606) I091D06 26-35 50-66  99-111 SRDLLLFPHSPLL(SEQ ID NO: 2456) I091D07 26-35 50-66  99-112 SRDLLLFPNGALRF(SEQ ID NO: 2645) I091D09 26-35 50-66  99-112 SRDLLLFPYSPLRF(SEQ ID NO: 2719) I091E01 26-35 50-66  99-112 SRDLLLFPDAPLHP(SEQ ID NO: 2425) I091E02 26-35 50-66  99-112 SRDLLLFPQAPLFP(SEQ ID NO: 2689) I091E03 26-35 50-66  99-112 SRDLLLFPSAPLWP(SEQ ID NO: 2352) I091E04 26-35 50-66  99-112 SRDLLLFPKSPLAF(SEQ ID NO: 2547) I091E06 26-35 50-66  99-112 SRDLLLFPSSPLHP(SEQ ID NO: 2576) I091E07 26-35 50-66  99-112 SRDLLLFPNHPLTF(SEQ ID NO: 2661) I091E08 26-35 50-66  99-112 SRDLLLFPNAPLDS(SEQ ID NO: 2607) I091E09 26-35 50-66  99-112 SRDLLLFPYAPLDF(SEQ ID NO: 2361) I091E10 26-35 50-66  99-112 SRDLLLFPSSPLEF(SEQ ID NO: 2711) I091F01 26-35 50-66  99-112 SRDLLLFPRAPLFF(SEQ ID NO: 2486) I091F03 26-35 50-66  99-112 SRDLLLFPMAPLVG(SEQ ID NO: 2599) I091F05 26-35 50-66  99-112 SRDLLLFPLAPLHP(SEQ ID NO: 2553) I091F06 26-35 50-66  99-112 SRDLLLFPHDPLGF(SEQ ID NO: 2353) I091F07 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I091F08 26-35 50-66  99-112 SRDLLLFPQSPLLF(SEQ ID NO: 2458) I091F09 26-35 50-66  99-112 SRDLLLFPHEHLSP(SEQ ID NO: 2354) I091F10 26-35 50-66  99-112 SRDLLLFPHSPLDF(SEQ ID NO: 2444) I091F11 26-35 50-66  99-112 SRDLLLFPHSPLSP(SEQ ID NO: 2549) I091F12 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I091G01 26-35 50-66  99-112 SRDLLLFPNAALYP(SEQ ID NO: 2386) I091G03 26-35 50-66  99-112 SRDLLLFPNDPLFG(SEQ ID NO: 2355) I091G04 26-35 50-66  99-112 SRDLLLFPGAPLSP(SEQ ID NO: 2478) I091G05 26-35 50-66  99-112 ARDLLLFPAAPLWP(SEQ ID NO: 2397) I091G06 26-35 50-66  99-111 SRDLLLFPNDPLR(SEQ ID NO: 2637) I091G07 26-35 50-66  99-112 SRDLLLFPTAPLDP(SEQ ID NO: 2345) I091G09 26-35 50-66  99-112 SRDLLLFPTAPLFP(SEQ ID NO: 2349) I091G10 26-35 50-66  99-112 SRDLLLFPSDPLVF(SEQ ID NO: 2660) I091G11 26-35 50-66  99-112 SRDLLLFPGSPLTF(SEQ ID NO: 2389) I091G12 26-35 50-66  99-112 SRDLLLFPYSHLEF(SEQ ID NO: 2655) I104A01 26-35 50-66  99-112 SRDLLLFPQSPLHP(SEQ ID NO: 2455) I104A07 26-35 50-66  99-112 SRDLLLFPQAPLFP(SEQ ID NO: 2689) I104A08 26-35 50-66  99-112 SRDLLLFPYAPLTF(SEQ ID NO: 2617) I104A09 26-35 50-66  99-112 SRDLLLFPQNPLHP(SEQ ID NO: 2506) I104A10 26-35 50-66  99-112 SRDLLLFPHEPLCF(SEQ ID NO: 2636) I104A11 26-35 50-66  99-112 SRDLLLFPSAPLSF(SEQ ID NO: 2611) I104Al2 26-35 50-66  99-112 SRDLLLFPMAPLRF(SEQ ID NO: 2593) I104B02 26-35 50-66  99-112 SRDLLLFPRSPLSF(SEQ ID NO: 2557) I104B04 26-35 50-66  99-112 SRDLLLFPSAPLYP(SEQ ID NO: 2387) I104B09 26-35 50-66  99-112 SRDLLLFPRDPLQF(SEQ ID NO: 2395) I104B11 26-35 50-66  99-112 SRDLLLFPTAPLTF(SEQ ID NO: 2531) I104C01 26-35 50-66  99-112 SRDLLLFPYSPLYP(SEQ ID NO: 2710) I104C04 26-35 50-66  99-112 SRDLLLFPASPLIF(SEQ ID NO: 2417) I104C05 26-35 50-66  99-112 SRDLLLFPRHPLLF(SEQ ID NO: 2543) I104C06 26-35 50-66  99-111 SRDLLLFPHAPLE(SEQ ID NO: 2524) I104C07 26-35 50-66  99-112 SRDLLLFPHAPLHP(SEQ ID NO: 2370) I104C09 26-35 50-66  99-112 SRDLLLFPHFPLIF(SEQ ID NO: 2399) I104C11 26-35 50-66  99-112 SRDLLLFPHEPLIF(SEQ ID NO: 2644) I104D01 26-35 50-66  99-112 SRDLLLFPNHAFDL(SEQ ID NO: 2652) I104D02 26-35 50-66  99-112 SRDLLLFPHTILYP(SEQ ID NO: 2497) I104D03 26-35 50-66  99-112 SRDLLLFPDWPLYP(SEQ ID NO: 2483) I104D04 26-35 50-66  99-112 SRDLLLFPHYPLFL(SEQ ID NO: 2511) I104D07 26-35 50-66  99-112 SRDLLLFPQAPLHP(SEQ ID NO: 2691) I104D08 26-35 50-66  99-112 SRDLLLFPHAPMDP(SEQ ID NO: 2595) I104D09 26-35 50-66  99-112 SRDLLLFPRAPLTF(SEQ ID NO: 2500) I104E01 26-35 50-66  99-112 SRDLLLFPRATLEF(SEQ ID NO: 2502) I104E02 26-35 50-66  99-112 SRDLLLFPHSPLFP(SEQ ID NO: 2447) I104E03 26-35 50-66  99-112 SRDLLLFPNDPLVL(SEQ ID NO: 2641) I104E05 26-35 50-66  99-112 SRDLLLFPHDPLYI(SEQ ID NO: 2463) I104E11 26-35 50-66  99-112 SRDLLLFPYAPLSF(SEQ ID NO: 2385) I104E12 26-35 50-66  99-112 SRDLLLFPASPLNP(SEQ ID NO: 2364) I104F02 26-35 50-66  99-112 SRDLLLFPHDPLSP(SEQ ID NO: 2616) I104F03 26-35 50-66  99-112 SRDLLLFPRDPLRF(SEQ ID NO: 2360) I104F04 26-35 50-66  99-112 SRDLLLFPGDPLDF(SEQ ID NO: 2481) I104F05 26-35 50-66  99-112 SRDLLLFPHGPLTF(SEQ ID NO: 2402) I104F06 26-35 50-66  99-112 SRDLLLFPHAPLSP(SEQ ID NO: 2573) I104F07 26-35 50-66  99-112 SRDLLLFPSSPLIL(SEQ ID NO: 2465) I104F10 26-35 50-66  99-112 SRDLLLFPNSPLSP(SEQ ID NO: 2362) I104F11 26-35 50-66  99-112 SRDLLLFPQDPLVF(SEQ ID NO: 2708) I104F12 26-35 50-66  99-112 SRDLLLFPKAPLVF(SEQ ID NO: 2544) I104G04 26-35 50-66  99-112 SRDLLLFPHAPLRF(SEQ ID NO: 2559) I104G05 26-35 50-66  99-112 SRDLLLFPRAPLAP(SEQ ID NO: 2476) I104G09 26-35 50-66  99-112 SRDLLLFPTAPLNF(SEQ ID NO: 2518) I104G11 26-35 50-66  99-112 SRDLLLFPQGPLSF(SEQ ID NO: 2482) I105A02 26-35 50-66  99-112 SRDLLLFPHLPLNP(SEQ ID NO: 2494) I105A03 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I105A04 26-35 50-66  99-112 SRDLLLFPGAPLAP(SEQ ID NO: 2487) I105A08 26-35 50-66  99-112 SRDLLLFPQAPLYP(SEQ ID NO: 2378) I105A09 26-35 50-66  99-112 SRDLLLFPRSPLSF(SEQ ID NO: 2557) I105A11 26-35 50-66  99-112 SRDLLLFPSHSFDI(SEQ ID NO: 2692) I105B04 26-35 50-66  99-112 SRDLLLFPYSPLHP(SEQ ID NO: 2658) I105B05 26-35 50-66  99-112 SRDLLLFPYSPLSF(SEQ ID NO: 2676) I105B07 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I105B08 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I105B10 26-35 50-66  99-112 SRDLLLFPASPLNP(SEQ ID NO: 2364) I105B11 26-35 50-66  99-112 SRDLLLFPHEPLSP(SEQ ID NO: 2651) I105B12 26-35 50-66  99-112 SRDLLLFPLDPLII(SEQ ID NO: 2560) I105C02 26-35 50-66  99-112 SRDLLLFPRAPLAF(SEQ ID NO: 2472) I105C03 26-35 50-66  99-112 SRDLLLFPSSPLSF(SEQ ID NO: 2715) I105C05 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I105C06 26-35 50-66  99-112 SRDLLLFPRAPLDF(SEQ ID NO: 2367) I105C08 26-35 50-66  99-112 SRDLLLFPRSPLTF(SEQ ID NO: 2562) I105C12 26-35 50-66  99-112 SRDLLLFPQHGFDA(SEQ ID NO: 2446) I105D04 26-35 50-66  99-112 SRDLLLFPRDPLRF(SEQ ID NO: 2360) I105D06 26-35 50-66  99-112 SRDLLLFPRDPLSF(SEQ ID NO: 2368) I105D08 26-35 50-66  99-112 SRDLLLFPYAPLAF(SEQ ID NO: 2608) I105D09 26-35 50-66  99-112 SRDLLLFPHAAFDV(SEQ ID NO: 2619) I105D10 26-35 50-66  99-112 SRDLLLFPHEPLFP(SEQ ID NO: 2640) I105D11 26-35 50-66  99-112 SRDLLLFPRSALTF(SEQ ID NO: 2519) I105E01 26-35 50-66  99-112 SRDLLLFPHHSFDS(SEQ ID NO: 2422) I105E06 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I105E11 26-35 50-66  99-112 SRDLLLFPNSPLHP(SEQ ID NO: 2675) I105F03 26-35 50-66  99-112 SRDLLLFPHHPLDS(SEQ ID NO: 2409) I105F06 26-35 50-66  99-112 SRDLLLFPQAPLHP(SEQ ID NO: 2691) I105F07 26-35 50-66  99-112 SRDLLLFPSWPLTF(SEQ ID NO: 2340) I105F09 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I105F12 26-35 50-66  99-112 SRDLLLFPTYPLVF(SEQ ID NO: 2604) I105G03 26-35 50-66  99-112 SRDLLLFPHAPLHP(SEQ ID NO: 2370) I105G08 26-35 50-66  99-112 SRDLLLFPKHPLVF(SEQ ID NO: 2366) I105G09 26-35 50-66  99-112 SRDLLLFPASPLNP(SEQ ID NO: 2364) I105G10 26-35 50-66  99-112 SRDLLLFPHHSFDA(SEQ ID NO: 2419) I105G11 26-35 50-66  99-112 SRDLLLFPHDPLLF(SEQ ID NO: 2614) I107A01 26-35 50-66  99-112 SRDLLLFPRHPLVF(SEQ ID NO: 2545) I107A03 26-35 50-66  99-112 SRDLLLFPRAPLYP(SEQ ID NO: 2501) I107A06 26-35 50-66  99-112 SRDLLLFPHAPLDP(SEQ ID NO: 2369) I107A07 26-35 50-66  99-112 SRDLLLFPNAPLSP(SEQ ID NO: 2371) I107A09 26-35 50-66  99-112 SRDLLLFPQAPLSP(SEQ ID NO: 2699) I107Al2 26-35 50-66  99-112 SRDLLLFPHAPLSF(SEQ ID NO: 2564) I107B02 26-35 50-66  99-112 SRDLLLFPHAPLFP(SEQ ID NO: 2533) I107B04 26-35 50-66  99-112 SRDLLLFPASPLTF(SEQ ID NO: 2420) I107B05 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I107C01 24-33 48-64  97-110 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I107C02 26-35 50-66  99-112 SRDLLLFPHYGMYV(SEQ ID NO: 2504) I107C04 26-35 50-66  99-112 SRDLLLFPHYPLHP(SEQ ID NO: 2357) I107C06 26-35 50-66  99-112 SRDLLLFPHAPLAP(SEQ ID NO: 2510) I107C08 26-35 50-66  99-112 SRDLLLFPQAPLEP(SEQ ID NO: 2681) I107C10 26-35 50-66  99-112 SRDLLLFPSHAFDL(SEQ ID NO: 2674) I107D01 26-35 50-66  99-112 SRDLLLFPYAPLDF(SEQ ID NO: 2361) I107D04 26-35 50-66  99-112 SRDLLLFPNAPLSF(SEQ ID NO: 2625) I107D07 26-35 50-66  99-112 SRDLLLFPSHSFDV(SEQ ID NO: 2693) I107D12 26-35 50-66  99-112 SRDLLLFPHHSFDT(SEQ ID NO: 2424) I107E01 26-35 50-66  99-112 SRDLLLFPMLGLDL(SEQ ID NO: 2499) I107E05 26-35 50-66  99-112 SRDLLLFPRAPLDF(SEQ ID NO: 2367) I107E07 26-35 50-66  99-112 SRDLLLFPRSPLLF(SEQ ID NO: 2551) I107E09 26-35 50-66  99-112 SRDLLLFPKAPLTF(SEQ ID NO: 2382) I107F01 26-35 50-66  99-112 SRDLLLFPSAPLSP(SEQ ID NO: 2623) I107F05 26-35 50-66  99-112 SRDLLLFPHAPLAP(SEQ ID NO: 2510) I107F09 26-35 50-66  99-112 SRDLLLFPSAPLAP(SEQ ID NO: 2394) I107F10 26-35 50-66  99-112 SRDLLLFPRTPLLF(SEQ ID NO: 2373) I107G01 26-35 50-66  99-112 SRDLLLFPNAPLSP(SEQ ID NO: 2371) I107G05 26-35 50-66  99-112 SRDLLLFPSAPLYP(SEQ ID NO: 2387) I107H02 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I107H06 26-35 50-66  99-112 SRDLLLFPRAPLSF(SEQ ID NO: 2496) I107H09 26-35 50-66  99-112 SRDLLLFPHYPLEM(SEQ ID NO: 2536) I107H10 26-35 50-66  99-112 SRDLLLFPHAPLAP(SEQ ID NO: 2510) I108Al2 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I108B03 26-35 50-66  99-112 SRDLLLFPRDPLLF(SEQ ID NO: 2515) I108B04 26-35 50-66  99-112 SRDLLLFPLSPLVP(SEQ ID NO: 2396) I108C09 26-35 50-66  99-112 SRDLLLFPHDPLGF(SEQ ID NO: 2353) I108C11 26-35 50-66  99-112 SRDLLLFPHHSLLF(SEQ ID NO: 2429) I108D10 26-35 50-66  99-112 SRDLLLFPASPLNP(SEQ ID NO: 2364) I108D11 26-35 50-66  99-112 SRDLLLFPASPLNP(SEQ ID NO: 2364) I108D12 26-35 50-66  99-112 SRDLLLFPSAPLNP(SEQ ID NO: 2709) I108E01 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I108E03 26-35 50-66  99-112 SRDLLLFPKHPLRF(SEQ ID NO: 2393) I108E05 26-35 50-66  99-112 SRDLLLFPHAPLFP(SEQ ID NO: 2533) I108E07 26-35 50-66  99-112 SRDLLLFPHAPLDP(SEQ ID NO: 2369) I108E08 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I108E09 26-35 50-66  99-112 SRDLLLFPSAPLSP(SEQ ID NO: 2623) I108E10 26-35 50-66  99-112 SRDLLLFPRDPLDL(SEQ ID NO: 2509) I108E11 26-35 50-66  99-112 SRDLLLFPRDPLEF(SEQ ID NO: 2516) I108F10 26-35 50-66  99-112 SRDLLLFPNAPLSP(SEQ ID NO: 2371) I108F12 26-35 50-66  99-112 SRDLLLFPHYPFDA(SEQ ID NO: 2508) I108G01 26-35 50-66  99-112 SRDLLLFPRDPLRF(SEQ ID NO: 2360) I108G02 26-35 50-66  99-112 SRDLLLFPDAPLAP(SEQ ID NO: 2381) I108G07 26-35 50-66  99-112 SRDLLLFPRAPLAP(SEQ ID NO: 2476) I108G10 26-35 50-66  99-112 SRDLLLFPHHSLLF(SEQ ID NO: 2429) I108G11 26-35 50-66  99-112 SRDLLLFPHHPLTF(SEQ ID NO: 2377) I108G12 26-35 50-66  99-112 SRDLLLFPHHPLTF(SEQ ID NO: 2377) I108H01 26-35 50-66  99-112 SRDLLLFPRDPLHF(SEQ ID NO: 2512) I108H02 26-35 50-66  99-112 SRDLLLFPNAPLNP(SEQ ID NO: 2615) I108H06 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I108H08 26-35 50-66  99-112 SRDLLLFPASPLNP(SEQ ID NO: 2364) I111A06 26-35 50-66  99-112 SRDLLLFPQAPLHP(SEQ ID NO: 2691) I111B12 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I111C01 26-35 50-66  99-112 SRDLLLFPQHGLDL(SEQ ID NO: 2449) I111D06 26-35 50-66  99-112 SRDLLLFPRDPLLF(SEQ ID NO: 2515) I111E04 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I111E10 26-35 50-66  99-112 SRDLLLFPQAPLHP(SEQ ID NO: 2691) I111E11 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I111E12 26-35 50-66  99-112 SRYLLLFPHHSFDL(SEQ ID NO: 2150) I111F07 26-35 50-66  99-112 SRDLLLFPRAPLYP(SEQ ID NO: 2501) I111G02 26-35 50-66  99-112 SRDLLLFPKAPLDF(SEQ ID NO: 2534) I111H10 26-35 50-66  99-112 SRYLLLFPQHGFDA(SEQ ID NO: 2703) I113A04 26-35 50-66  99-112 SRDLLLFPSAPLWP(SEQ ID NO: 2352) I113Al2 26-35 50-66  99-112 SRDLLLFPQEPLAP(SEQ ID NO: 2434) I113B06 26-35 50-66  99-112 SRDLLLFPHHPLEP(SEQ ID NO: 2411) I113C06 26-35 50-66  99-112 SRDLLLFPHHGFDA(SEQ ID NO: 2406) I113G04 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I113G05 26-35 50-66  99-112 SRDLLLFPHYSLLL(SEQ ID NO: 2517) I113G10 26-35 50-66  99-112 SRDLLLFPHHPLQF(SEQ ID NO: 2413) I113G11 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I113H06 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I113H07 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I113H09 26-35 50-66  99-112 SRDLLLFPHYTLLF(SEQ ID NO: 2525) I114C04 26-35 50-66  99-112 SRDLLLFPHHGFDA(SEQ ID NO: 2406) I114C12 26-35 50-66  99-112 SRDLLLFPQAPLHP(SEQ ID NO: 2691) I114D04 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I114D06 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I114D10 26-35 50-66  99-112 SRDLLLFPHYSLVL(SEQ ID NO: 2521) I114E01 26-35 50-66  99-112 SRDLLLFPQEPLSP(SEQ ID NO: 2435) I114E02 26-35 50-66  99-112 SRDLLLFPQESFSL(SEQ ID NO: 2437) I114E03 26-35 50-66  99-112 SRDLLLFPKAPLTF(SEQ ID NO: 2382) I114E11 26-35 50-66  99-112 SRDLLLFPHDSFFL(SEQ ID NO: 2383) I114H01 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I114H06 26-35 50-66  99-112 SRDLLLFPHHALDV(SEQ ID NO: 2404) I114H09 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I115A02 26-35 50-66  99-112 SRYLLLFPDHSFDL(SEQ ID NO: 2684) I115A07 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I115B10 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I115C05 26-35 50-66  99-112 SRDLLLFPRAPLYP(SEQ ID NO: 2501) I115C06 26-35 50-66  99-112 SRYLLLFPHHSFDL(SEQ ID NO: 2150) I115C08 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I115C12 26-35 50-66  99-112 SRDLLLFPHHSFDT(SEQ ID NO: 2424) I115D07 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I115E09 26-35 50-66  99-112 SRDLLLFPHHRFDL(SEQ ID NO: 2418) I115F06 26-35 50-66  99-112 SRYLLLFPHYGMDV(SEQ ID NO: 2685) I115F07 26-35 50-66  99-112 SRYLLLFPHYPLLF(SEQ ID NO: 2686) I115F12 26-35 50-66  99-112 SRYLLLFPHHSFDL(SEQ ID NO: 2150) I115G04 26-35 50-66  99-112 SRDLLLFPHHRFDL(SEQ ID NO: 2418) I115G05 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I115G08 26-35 50-66  99-112 SRDLLLFPHDSFDL(SEQ ID NO: 2631) I115H04 26-35 50-66  99-112 SRDLLLFPHANLSP(SEQ ID NO: 2503) I115H07 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I115H09 26-35 50-66  99-112 SRDLLLFPHHRFDL(SEQ ID NO: 2418) I116A07 26-35 50-66  99-112 SRDLLLFPYEPLRF(SEQ ID NO: 2642) I116B01 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I116B12 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I116C06 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I116D07 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I116E02 26-35 50-66  99-112 SRDLLLFPHHRFDL(SEQ ID NO: 2418) I116E04 26-35 50-66  99-112 SRDLLLFPHHSFDL(SEQ ID NO: 2147) I116F02 26-35 50-66  99-112 SRYLLLFPHHSFDL(SEQ ID NO: 2150) I116F11 26-35 50-66  99-112 SRDLLLFPHYPLLF(SEQ ID NO: 2344) I116G05 26-35 50-66  99-112 SRDLLLFPQAPLSP(SEQ ID NO: 2699) I001C09 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I006D07 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I007B03 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I007F11 26-35 50-66  99-113 DGIDILLVPAALMDV(SEQ ID NO: 2160) I007H08 26-37 52-67 102-117 DRYDILTGYYYYGMDV(SEQ ID NO: 2129) I008A09 26-35 50-66  99-119 DREAYYDILTGYYLYYYYMDV(SEQ ID NO: 2172) I008B01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I008C02 26-37 52-67 100-118 HVRDYDILTGYYRGHYFDY(SEQ ID NO: 2167) I008C03 26-35 50-65  98-116 EGSYDILTGYYVGVGRMDV(SEQ ID NO: 2171) I008C12 26-35 50-68 101-119 FNPTYDILTGYYIGGYFQH(SEQ ID NO: 2155) I012A06 26-37 52-67 100-118 GRWDYDLLTGEHLGYYFDY(SEQ ID NO: 2162) I016E05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016F02 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I016F04 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016H07 26-35 50-66  99-114 GYHDPLTSYNYNWFDP(SEQ ID NO: 2163) I018C02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018C10 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I018D07 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I018H08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018H09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I021B05 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I022E02 26-35 50-66  99-114 ATYDPLTGYSFDGLDI(SEQ ID NO: 2157) I026E03 26-35 50-66  99-114 TDYDILTGYPMGYFDP(SEQ ID NO: 2173) I027A07 26-35 50-66  99-117 GGEYDILTGYYFGLGVYDY(SEQ ID NO: 2170) I028A06 26-35 50-66  99-115 GGDYDILTGLYYYGMDV(SEQ ID NO: 2156) I029D07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I029F11 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I031C03 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I031C07 26-35 50-66  99-120 SSPPRWYDALTGDSSYHSAMDV(SEQ ID NO: 2169) I031F09 26-35 50-66  99-116 DEGRDLLTGYYWPNFFDS(SEQ ID NO: 2168) I031G08 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I031G10 26-35 50-66  99-120 SSPPKWYDALTGDSSYHSAMDV(SEQ ID NO: 2165) I031G11 26-35 50-66  99-116 DEGRDLLTGYYWPNFFDS(SEQ ID NO: 2168) I037E07 26-35 50-66  99-113 DGIDILLVPAALMDV(SEQ ID NO: 2160) I037E12 26-35 50-66  99-113 DGIDILLVPAALMDV(SEQ ID NO: 2160) I050A07 26-40 55-71 104-118 QDNDPLTGYKLGFDY(SEQ ID NO: 2164) I061D02 26-37 52-69 102-117 DRYDILTGYYYYGMDV(SEQ ID NO: 2129) I061E07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I061H01 26-35 50-68 101-119 FNPTYDILTGYYIGGYFQH(SEQ ID NO: 2155) I001A03 26-35 50-66  99-117 ERHYYDILTGYQTGYGMDV(SEQ ID NO: 2784) I001A07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001A08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001A10 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001Al2 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001B02 26-35 50-66  99-110 DRETKVGYGMDV(SEQ ID NO: 2945) I001B07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001C06 24-33 49-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I001C08 26-35 50-66  99-117 EGSYDILTGYYVGVGRMDV(SEQ ID NO: 2171) I001C12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001D08 26-35 50-66  98-113 DSYDILTGYRGYYFDY(SEQ ID NO: 2745) I001D12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001E05 24-33 49-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I001E07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001G09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I001H05 26-35 50-66  99-117 ERHYYDILTGYQTGYGMDV(SEQ ID NO: 2784) I001H08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I003A01 26-34 49-65  98-113 ELGSSIVGATTGALDM(SEQ ID NO: 2852) I003A06 26-34 49-65  98-113 ELGSSIVGATTGALDM(SEQ ID NO: 2852) I003A07 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I003A10 26-35 50-66  99-114 MEYDILTGYYGGYFDY(SEQ ID NO: 2179) I003B03 26-34 49-65  98-113 ELGSSIVGATTGALDM(SEQ ID NO: 2852) I003B04 25-34 49-65  98-111 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I003B09 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I003C01 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I003C02 26-35 50-66  99-114 GDYDILTGYPAECFQI(SEQ ID NO: 2854) I003C03 26-35 50-66  99-114 GDYDILTGYPAECFQI(SEQ ID NO: 2854) I003C12 26-35 50-66  99-114 MEYDILTGYYGGYFDY(SEQ ID NO: 2179) I003D04 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I003E05 26-35 50-66  99-114 GDYDILTGYPAECFQI(SEQ ID NO: 2854) I003F01 26-34 49-65  98-113 ELGSSIVGATTGALDM(SEQ ID NO: 2852) I003F02 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I003G01 26-35 50-66  99-116 GTGYDILTGYYMGSAFDQ(SEQ ID NO: 2800) I003G05 26-35 50-66  99-116 GSGYDLLTGYFTGSPLDY(SEQ ID NO: 2766) I003G06 26-35 50-66  99-118 DRGGNYDILTGYYFHHGVDV(SEQ ID NO: 2914) I003G11 26-35 50-66  99-117 DAQSYYDILTGYQSYAFDI(SEQ ID NO: 2183) I003H02 26-35 50-66  98-113 DNYDILTGYSRRFDP(SEQ ID NO: 2942) I003H05 26-34 49-65  98-113 ELGSSIVGATTGALDM(SEQ ID NO: 2852) I003H08 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I005A01 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I005A02 26-35 50-66  99-114 EGRDILTGVYYYGLDV(SEQ ID NO: 2893) I005B01 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I005B09 26-35 50-66  98-110 TYYDILTGRFFDI(SEQ ID NO: 2866) I005C01 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I005D02 26-35 50-66  99-114 DLRYDILTGYHDAFDI(SEQ ID NO: 2890) I005D03 26-35 50-66  99-115 GAYYDILTGYYPYGMDV(SEQ ID NO: 2860) I005E01 26-35 50-66  99-115 GTYYDILTGYFHYGMDV(SEQ ID NO: 2774) I005E08 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I005F01 26-35 50-66  99-113 DQHDILTGVYYGMDV(SEQ ID NO: 2921) I005F02 26-35 50-66  99-117 VSPSYDILTGYYLPHAFDV(SEQ ID NO: 2849) I005F04 26-35 50-65  98-110 TYYDILTGRFFDI(SEQ ID NO: 2866) I005F08 26-35 50-66  99-113 PSYDILTGYLYYFDY(SEQ ID NO: 2850) I005G01 26-35 50-66  99-114 DLRYDILTGYHDAFDI(SEQ ID NO: 2890) I005G08 26-35 50-66  99-115 GAYYDILTGYYPYGMDV(SEQ ID NO: 2860) I005H02 26-35 50-66  99-113 GQYYDILTGYNWFDP(SEQ ID NO: 2857) I006B01 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I006C09 26-35 50-66  99-116 GGYSSGWLRGGPYNWFDP(SEQ ID NO: 2967) I006D09 26-35 50-66  99-114 GDYDILTGYYIPLRDY(SEQ ID NO: 2792) I006E01 26-35 50-68 101-116 NLFDVWTLPYYYYMDV(SEQ ID NO: 2965) I006E07 26-35 50-66  99-116 ADYDILTGYSPLTYGMDV(SEQ ID NO: 2762) I006F01 26-35 50-68 101-113 MYYDILTGHNFDY(SEQ ID NO: 2879) I006F02 26-35 50-66  99-115 VSRDILTGNYYYYGMDV(SEQ ID NO: 2817) I006F07 26-35 50-66  99-116 GGYSSGWLRGGPYNWFDP(SEQ ID NO: 2967) I006G01 26-35 50-68 101-119 AGGYYDILTGRDYYYGMDV(SEQ ID NO: 2877) I006G04 26-35 50-66  99-105 RRYALDY (SEQ ID NO: 2920)I006H01 26-35 50-65  98-119 DRGSYDILTGYYTPPHYYGMDV (SEQ ID NO: 2761)I006H02 26-35 50-66  99-116 GGYSSGWLRGGPYNWFDP (SEQ ID NO: 2967) I007A0126-35 50-66  99-114 ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I007A08 26-3550-66  99-114 SHYDILTGLNYWYFDY (SEQ ID NO: 2746) I007A11 26-35 50-66 99-113 ENYDFLTGYYGAFDI (SEQ ID NO: 2772) I007Al2 26-35 50-68 101-117GIYDILTGYHWDGAFDI (SEQ ID NO: 2892) I007B04 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I007C04 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I007C08 26-35 50-65  98-115IRLYCYSLTGYYPYGMDD (SEQ ID NO: 2810) I007C12 26-35 50-66  99-113TNYDILTGYYQGVDY (SEQ ID NO: 2782) I007D07 26-35 50-66  99-113GQYYDILTGYNWFDP (SEQ ID NO: 2857) I007D08 26-35 50-68 101-117GIYDILTGYHWDDAFDI (SEQ ID NO: 2872) I007E03 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I007E10 26-35 50-66  99-114DFYDILTGYPLGGMDV (SEQ ID NO: 2741) I007E11 26-35 50-66  99-117DLPYYDILTGYSLTSGMDV (SEQ ID NO: 2923) I007F06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I007F08 26-35 50-65  98-116GRRYDILTGYYYYHHGMDV (SEQ ID NO: 2811) I007G07 26-35 50-66  99-114SHYDILTGLNYWYFDL (SEQ ID NO: 2166) I007G09 26-35 50-66  99-115DSGGDILTGYYMPYFDY (SEQ ID NO: 2847) I007G10 26-35 50-65  98-115VGLYYDILTGYYPSGMDV (SEQ ID NO: 2805) I007H07 26-35 50-68 101-120SQAHYDILTGYYLWSYGMDV (SEQ ID NO: 2875) I007H11 26-35 50-66  99-114ESYDILTGYRHYGMDL (SEQ ID NO: 2891) I008A02 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008A05 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008A06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008A07 26-35 50-66  99-115DREYDLLTGYYLHAFDM (SEQ ID NO: 2960) I008Al2 26-35 50-66  99-113ENYDFLTGYYGAFDI (SEQ ID NO: 2772) I008B02 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008B04 26-35 50-66  99-116DGSYDILTGYYIDNYMDV (SEQ ID NO: 2154) I008B05 26-35 50-66  99-114DHYDILTGLYYYGMDV (SEQ ID NO: 2760) I008B06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008B07 24-33 48-64  97-113GRRYDILTGYYKGPLDY (SEQ ID NO: 2902) I008B10 26-35 50-66  99-114AYYDNLTGFLPYGMGV (SEQ ID NO: 2947) I008B11 26-35 50-66  99-117EGYDILTGYFLDYYHGMDV (SEQ ID NO: 2753) I008C06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008C08 26-35 50-66  99-122GPRGGPYYDILTGYYLSLSDAFDI (SEQ ID NO: 2729) I008C09 26-35 50-66  99-115EYYDILTGYRDPYGMDV (SEQ ID NO: 2973) I008D01 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008D02 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008D03 26-35 50-66  99-117EVRNYDLLTRSYLAGPLDN (SEQ ID NO: 2751) I008D04 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008D05 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008D06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008D07 26-35 50-66  99-117DRGYYDILTGYYRGHGMDV (SEQ ID NO: 2837) I008D08 26-35 50-66  99-117DLPYYDILTGYSLTSGMDV (SEQ ID NO: 2923) I008D12 26-35 50-66  99-117EEGFYDILTGYYGPGYFDY (SEQ ID NO: 2974) I008E01 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008E02 20-31 46-63  96-110EGYDILTGYSKFLDY (SEQ ID NO: 2906) I008E03 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008E04 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008E08 26-35 50-66  99-114SHYDILTGLNYWYFDL (SEQ ID NO: 2166) I008E09 26-35 50-66  99-116ERADYDILTGYYFYDMDV (SEQ ID NO: 2833) I008E12 26-37 52-67 100-114FRYDILTSYYYGMDV (SEQ ID NO: 2734) I008F03 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008F06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I008F07 26-35 50-65  98-116GRRYDILTGYYYYHHGMDV (SEQ ID NO: 2811) I008F08 26-35 50-66  99-116GHYDILTGYDDYYYGMDV (SEQ ID NO: 2844) I008F09 26-35 50-65  98-106HDILTGFDY (SEQ ID NO: 2904) I008F10 26-35 50-66  99-113 SGYDILTGYLYGMDV(SEQ ID NO: 2934) I008F11 26-35 50-68 101-117 APYDILTGYSDYYGMDV(SEQ ID NO: 2968) I008G02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I008G03 26-35 50-66  99-113 GDYDPLTGYSFGVDV(SEQ ID NO: 2941) I008G04 26-35 50-65  98-116 EGSYDILTGYYVGVGRMDV(SEQ ID NO: 2171) I008G05 26-35 50-66  99-117 DGYYDILTGGFYYYYGMDV(SEQ ID NO: 2899) I008G11 26-35 50-66  99-109 AYYDILTGLDY(SEQ ID NO: 2966) I008G12 26-35 50-66  99-116 DQQYDILTGYYIHYGMDV(SEQ ID NO: 2964) I008H02 26-35 50-66  99-114 DQVDLLLMDHNYYMDV(SEQ ID NO: 2918) I008H03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I008H06 26-35 50-66  98-116 EGSYDILTGYYVGVGRMDV(SEQ ID NO: 2171) I008H09 26-35 50-66  99-116 DQQYDILTGYYIHYGMDV(SEQ ID NO: 2964) I008H11 26-35 50-66  99-114 TKYDILTGYYYYYMDV(SEQ ID NO: 2856) I012B03 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012B06 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012B10 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012C03 26-35 50-66  99-115 TDRFGAKDVTSRWGMDV(SEQ ID NO: 2814) I012C06 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012C09 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012D12 26-35 50-66  99-118 DRGGNYDILTGYYFHHGVDV(SEQ ID NO: 2914) I012E07 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012E08 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I012E09 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012F05 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012F12 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012G03 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I012G05 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I012G10 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I012H09 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I013A10 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I013Al2 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I013B04 26-35 50-66  99-120 SSPPKWYDALTGDSSYHSAMDV(SEQ ID NO: 2165) I013B09 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I013C02 26-35 50-66  99-120 SSPPKWYDALTGDSSYRSAMDV(SEQ ID NO: 2818) I013C04 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I013D02 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I013D03 26-35 50-66  99-120 SSPPKWYDALTGDSSYHSAMDV(SEQ ID NO: 2165) I013D10 26-35 50-66  99-118 GLRHVTLFGTGTRGHFYMDV(SEQ ID NO: 2789) I013E02 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I013E05 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I013E09 26-35 50-66  99-120 SSPPKWYDALTGDSSYHSAMDV(SEQ ID NO: 2165) I013F03 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I013F04 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I013F07 26-35 50-66  99-118 AATTSQKHNKYAYYFYGMDV(SEQ ID NO: 2131) I013F09 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I013F10 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I013H04 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I013H07 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I014Al2 24-33 49-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I014C06 26-35 50-66  99-114 GDYDILTGYPAECFQI(SEQ ID NO: 2854) I014C10 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I014C12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I014E06 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I014F02 26-37 52-67 100-114 AGYDLLTGYPFYFDS(SEQ ID NO: 2757) I016A08 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I016A09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016C02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016C03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016C05 26-35 50-66  99-121 VQMDSEYYDLLTGINVGPYYFDY(SEQ ID NO: 2132) I016C09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016C11 26-35 50-66  99-121 VQMDSEYYDLLTGINVGPYYFDY(SEQ ID NO: 2132) I016D10 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016D11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016E03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016E04 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016F03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016F11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016G01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016G06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I016G12 26-35 50-66  99-121 VQMDSEYYDLLTGINVGPYYFDY(SEQ ID NO: 2132) I016H10 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I017A06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I017A07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I017A11 25-34 49-65  98-113 ATYDPLTGYSFDGFDI(SEQ ID NO: 2157) I017E12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I017G03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I017G07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I017H01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018A02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018A04 26-35 50-66  99-117 EGSYDILTGYYVGVGRMDV(SEQ ID NO: 2171) I018A05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018A11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018B02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018B08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018C04 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018D02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018E06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018E08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018F04 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018G06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I018H07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I019E05 26-35 50-66  99-117 ERHYYDILTGYQTGYGMDV(SEQ ID NO: 2784) I019F06 26-35 50-66  99-117 ERHYYDILTGYQTGYGMDV(SEQ ID NO: 2784) I019G12 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I020D01 26-35 50-66  99-110 DRETKVGYGMDV(SEQ ID NO: 2945) I020D05 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I020E10 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I020G12 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I020H06 24-33 48-64  97-116 EGGNYHILTGYYIGNGAFDI(SEQ ID NO: 2896) I020H10 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2903) I021A11 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I021B01 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I021C11 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I021D12 26-35 50-66  99-110 DRETKVGYGMDV(SEQ ID NO: 2945) I021E10 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I021G02 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I022A08 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I022B01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I022B10 26-35 50-66  99-114 MEYDILTGYYGGYFDY(SEQ ID NO: 2179) I022C02 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I022C04 26-35 50-66  99-114 ATYDPLETGYSFDGFDI(SEQ ID NO: 2153) I022C08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I022D06 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I022E08 26-35 50-66  99-115 ASYYDILTGYYKGAFDI(SEQ ID NO: 2855) I022F01 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I022F04 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I022F12 26-35 50-66  99-113 GDYDILTGTYYYIDV(SEQ ID NO: 2859) I022G11 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I023D01 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I023D04 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I024B04 26-35 50-66  99-113 VYYDILTGYNLFFDY(SEQ ID NO: 2177) I024D01 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I024F06 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I024H01 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I024H07 26-35 50-66  99-115 DGYYDILTGYSYYGMDV(SEQ ID NO: 2135) I025A01 26-34 49-65  98-113 ELGSSIVGATTGALDM(SEQ ID NO: 2852) I025A04 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I025A07 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I025B01 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I025B10 26-35 50-66  98-113 DNYDILTGYSRRFDP (SEQ ID NO: 2942) I025B1226-34 49-65  98-113 ELGSSIVGATTGALDM (SEQ ID NO: 2852) I025C07 26-3449-65  98-113 ELGSSIVGATTGALDM (SEQ ID NO: 2852) I025D11 26-34 49-65 98-113 ELGSSIVGATTGALDM (SEQ ID NO: 2174) I025E04 26-35 50-66  99-115PLGITAVRGAKTDAFGI (SEQ ID NO: 2929) I025E05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I025E07 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I025E10 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I025F01 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I025F08 26-35 50-66  99-110GGSSQNFYGMDV (SEQ ID NO: 2884) I025G03 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I025G08 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I025H02 26-35 50-65  98-117AGSGFHDILTGYYKGGYFDY (SEQ ID NO: 2961) I026A01 26-35 50-66  99-114GDYDILTGYPAECFQI (SEQ ID NO: 2854) I026B01 26-35 50-66  99-116GSVYDILTGTYYKSGMGV (SEQ ID NO: 2733) I026B06 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026C06 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026C10 26-34 49-65  98-111RYGDPFYYYYYMNV (SEQ ID NO: 2755) I026C11 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I026D09 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I026E04 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I026E06 26-35 50-66  99-113GYDDILTGYIMALDY (SEQ ID NO: 2821) I026E09 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026F01 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026F09 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026F12 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026G08 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026G10 26-35 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I026G11 26-35 50-66  99-116GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I026H02 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I026H06 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I026H10 26-35 50-66  99-117GGEYDILTGYYFGLGVYDY (SEQ ID NO: 2170) I027A09 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I027B02 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I027B05 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I027C08 26-34 49-63  96-111ELGSSIVGATTGALDM (SEQ ID NO: 2852) I027D02 26-35 50-66  99-114DPFGAVPGYYYYAMDV (SEQ ID NO: 2826) I027E03 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I027E05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I027F04 26-35 50-66  99-117GPWYDPLFPPSGRHYGLDV (SEQ ID NO: 2793) I027F05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I027F11 26-34 49-65  98-113ELGSSIVGATTGALDM (SEQ ID NO: 2852) I027G06 26-35 50-66  99-113DNYDILTGYSRRFDP (SEQ ID NO: 2942) I027G07 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I027H03 26-35 50-66  99-114GDYDILTGYPAECFQI (SEQ ID NO: 2854) I028A04 26-35 50-66  99-116DMYYDILTGYYTGLAFDM (SEQ ID NO: 2880) I028A07 26-35 50-66  99-114VLNYDILTGYYYGMDV (SEQ ID NO: 2832) I028B08 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I028B10 26-35 50-68 101-121DFGYYDILTGYYIGAFYAFDI (SEQ ID NO: 2861) I028C01 26-37 52-69 102-115GGHTCIIPTCHMGG (SEQ ID NO: 2796) I028C04 26-35 50-66  99-116DMYYDILTGYYTGLAFDM (SEQ ID NO: 2880) I028C08 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I028D04 26-35 50-65  98-113ATQDILTGYLYSGMDV (SEQ ID NO: 2977) I028D05 26-35 50-66  99-114EHYDILTGYSLLGMDV (SEQ ID NO: 2907) I028D12 26-35 50-66  99-116DGYYDILTGYSVYYGMDV (SEQ ID NO: 2938) I028E06 26-35 50-65  98-116EGSYDILTGYYVGVGRMDV (SEQ ID NO: 2171) I028E07 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I028E08 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I028F06 26-35 50-66  99-119DDRRGYYDILTGYYRFGSFDI (SEQ ID NO: 2901) I028F08 26-35 50-66  99-107DIDIGGDDS (SEQ ID NO: 2954) I028G08 26-35 50-66  99-114 VSGYNSGYFESYDMDV(SEQ ID NO: 2732) I028G09 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I028G10 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I028H02 26-37 52-69 102-115 SGEPCITLACNLGG(SEQ ID NO: 2797) I028H03 26-35 50-66  99-121 DASEYYDILTGYYLATGRNWFDP(SEQ ID NO: 2888) I028H06 26-35 50-66  99-118 DPSYYDILTGYFLPYYMDV(SEQ ID NO: 2843) I028H09 26-35 50-68 101-113 EISSILTGYYMDV(SEQ ID NO: 2905) I029A10 26-35 50-65  98-112 MNYDILTGLVNWFDP(SEQ ID NO: 2786) I029Al2 26-35 50-68 101-110 RDILTGFYDS(SEQ ID NO: 2933) I029B11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I029C08 26-35 50-66  99-117 EGSYDILTGYYVGVGRMDV(SEQ ID NO: 2171) I029E10 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I029F08 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I029G08 26-35 50-66  99-114 GYYDILTGYQSDAFDI(SEQ ID NO: 2927) I030A02 26-35 50-66  99-115 TERFGAKDVTARWGMDV(SEQ ID NO: 2874) I030A03 26-35 50-66  99-113 ENYDILTGYYNFFDY(SEQ ID NO: 2737) I030A04 26-35 50-66  99-113 RQYDILTGYYGGFDY(SEQ ID NO: 2958) I030A05 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I030A09 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I030Al2 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I030B06 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I030B08 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I030B10 26-35 50-66  99-114 ELGHREGGYWYSPYNV(SEQ ID NO: 2838) I030003 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I030006 26-35 50-68 101-119 DPGNYDILTGYYYYYGMDV(SEQ ID NO: 2935) I030008 26-35 50-66  99-106 SGPGWFDP (SEQ ID NO: 2870)I030009 26-34 49-65  98-113 ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030C1026-34 49-65  98-113 ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030C11 26-3550-66  99-112 RYGDPFYYYYYMNV (SEQ ID NO: 2755) I030C12 26-35 50-66 99-106 SGPGWFDP (SEQ ID NO: 2870) I030D07 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I030D12 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030E02 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I030E05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030E07 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030E08 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030E09 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030E10 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I030F02 26-37 52-67 100-114AGYDLLTGYPFYFDS (SEQ ID NO: 2757) I030F05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030F06 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I030F08 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030F09 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030F11 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I030F12 26-35 50-66  99-113DNYDILTGYSRRFDP (SEQ ID NO: 2942) I030G03 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030G07 26-35 50-66  99-112RYGDPFYYYYMNV (SEQ ID NO: 2755) I030G09 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I030H05 26-35 50-66  99-118DRGGNYDILTGYYFHHGVDV (SEQ ID NO: 2914) I030H06 26-37 52-69 102-116ATKSYDILTRMYYYHMDV (SEQ ID NO: 2748) I030H10 26-35 50-66  99-113DNYDILTGYSRRFDP (SEQ ID NO: 2942) I030H11 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I031A01 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031A03 26-35 50-66  99-114PYYDPLTAYTFQYFGN (SEQ ID NO: 2806) I031A08 26-35 50-66  99-120GREDTDKVKPWDRYYHYYYMDV (SEQ ID NO: 2809) I031Al2 26-35 50-66  99-119GREDTDKVKPWDRYYHYYMDV (SEQ ID NO: 2972) I031B03 26-35 50-68 101-109GLGHTDSDS (SEQ ID NO: 2959) I031B06 26-35 50-66  99-115AKGYYYDSSGASDVFDV (SEQ ID NO: 2871) I031B07 26-35 50-66  99-120GREDTDKVKPWDRYYHYYYMDV (SEQ ID NO: 2809) I031B08 26-35 50-66  99-120SSPPKWYDALTGHSSYHSAMDV (SEQ ID NO: 2159) I031B09 26-35 50-66  99-120SNPPKWYDALTGHSSYHSAMDV (SEQ ID NO: 2840) I031B11 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031B12 26-35 50-66  99-120GREDTDKVKPWDRYYHYYYMDV (SEQ ID NO: 2809) I031C01 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031C02 26-35 50-66  99-114PFYDTLTSYVFQYFDH (SEQ ID NO: 2137) I031C04 26-35 50-66  99-120GRKDTDKVKPWDRYYHYYYMDV (SEQ ID NO: 2813) I031C08 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031C11 26-35 50-66  99-120GREDTDKVKPWDRYYHYYYMDV (SEQ ID NO: 2809) I031D01 26-35 50-66  99-118AATTSQKHNKYAYYFYGMDV (SEQ ID NO: 2131) I031D04 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031D06 26-35 50-66  99-120GREDTDKVKLWDRYYHYYYMDV (SEQ ID NO: 2807) I031D08 26-35 50-66  99-117VRPKLRYFDWLSRHDAFDL (SEQ ID NO: 2820) I031D09 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031D11 26-35 50-66  99-120SSPPKWYDALTGDSSYHSAMDV (SEQ ID NO: 2165) I031D12 26-35 50-66  99-117DKAHGEYGRDYYYYYGMDV (SEQ ID NO: 2735) I031E01 26-35 50-66  99-120SSPPKWYDALTGHSSYHSAMDV (SEQ ID NO: 2159) I031E05 26-35 50-66  99-120SSPPKWYDALTGHSSYHSAMDV (SEQ ID NO: 2848) I031E07 26-35 50-66  99-120SSPPKWYDALTGHSSYHSAMDV (SEQ ID NO: 2159) I031E08 26-35 50-66  99-120GREDTDKVKPWDRYYHYYYMDV (SEQ ID NO: 2809) I031E09 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031E10 26-35 50-66  99-120SSPPKWYDALTGDSSYHSAMDV (SEQ ID NO: 2165) I031E11 26-35 50-66  99-120SSPPKWYDALTGHSSYHSAMDV (SEQ ID NO: 2159) I031F01 26-35 50-66  99-110GYDSSAFRAFDI (SEQ ID NO: 2136) I031F04 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I031F06 26-35 50-66  99-108 DTVRSGGMDV(SEQ ID NO: 2804) I031F10 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I031F11 26-35 50-66  99-117 DKAHGEYGRDYYYYYGMDV(SEQ ID NO: 2735) I031F12 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I031G01 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I031G03 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I031G05 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I031G06 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I031G07 26-35 50-66  99-120 SSPPKWYDALTGDSSYHSAMGV(SEQ ID NO: 2816) I031G09 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I031G12 26-35 50-66  99-118 AATTSQKHNKYAYYFYGMDV(SEQ ID NO: 2131) I031H01 26-35 50-66  99-110 GYDSSAFRAFDI(SEQ ID NO: 2136) I031H02 26-35 50-66  99-115 AKGYYYDSSGASDVFDV(SEQ ID NO: 2871) I031H03 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I031H06 26-35 50-66  99-117 DKAHGEYGRDYYYYYGMDV(SEQ ID NO: 2735) I031H09 26-35 50-66  99-117 DKAHGEYGRDYYYYYGMDV(SEQ ID NO: 2735) I031H10 26-35 50-66  99-116 DRGYTGYDRLVGGYYFDF(SEQ ID NO: 2931) I031H11 26-35 50-66  99-108 DTVRSGGMDV(SEQ ID NO: 2804) I033A08 26-37 50-66 102-117 DRYDILTGYYYYGMDV(SEQ ID NO: 2129) I033B11 26-37 50-66 102-117 DRYDILTGYYYYGMDV(SEQ ID NO: 2129) I033C01 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I033C08 26-35 50-66  99-115 EMGYDILTGYYLNYMDV(SEQ ID NO: 2862) I033D02 26-35 50-66  99-111 GDYDILTGYYMDV(SEQ ID NO: 2781) I033D03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I033D05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I033D11 26-35 50-66  99-113 VKRDILTGYVEGMDV(SEQ ID NO: 2869) I033D12 26-35 50-66  99-117 GGPHYDILTGYYMAVGFDI(SEQ ID NO: 2962) I033E01 26-35 50-66  99-112 DIDARLAALDAFDI(SEQ ID NO: 2794) I033E06 26-35 50-66  99-114 ATHDPLTGYSFDGFDI(SEQ ID NO: 2780) I033E11 26-35 50-66  99-116 HRSRSCSSTSCRNDAFDI(SEQ ID NO: 2770) I033E12 26-35 50-66  99-115 EMGYDILTGYYLNYMDV(SEQ ID NO: 2862) I033F03 26-35 50-66  99-112 EGAADYLNGQYFQD(SEQ ID NO: 2768) I033F08 26-35 50-66  99-118 QKVYYDILTGYNYYYYGMDV(SEQ ID NO: 2767) I033F10 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I033F12 26-35 50-66  99-107 DIDIGGDDS(SEQ ID NO: 2954) I033G01 24-33 48-64  97-116 EGGNYDILTGYYIGNGAFDI(SEQ ID NO: 2158) I033G03 26-35 50-66  99-115 PQGVTLVRGAETDAFAI(SEQ ID NO: 2925) I033G08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I033H04 25-34 49-65  98-113 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I037A05 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I037B03 26-35 50-66  99-114 SHYDILTRLNYWYFDL(SEQ ID NO: 2950) I037B04 26-35 50-66  99-117 DPGYYDILTGYFHRYGMDV(SEQ ID NO: 2922) I037C04 26-35 50-65  98-115 ENGDYDILTGQTFYGMDV(SEQ ID NO: 2752) I037C06 26-35 50-66  99-114 LYYDILTGYHWDAFDI(SEQ ID NO: 2882) I037C08 26-35 50-66  99-113 DGIDILLVPAALMDV(SEQ ID NO: 2160) I037D11 26-35 50-66  99-109 SQWLEHDVFDI(SEQ ID NO: 2864) I037E06 26-35 50-66  99-117 DRRDYDLLTRYYYYYGMDV(SEQ ID NO: 2928) I037F04 26-35 50-65  98-117 KQRGDYDILTGYQLGYAFDI(SEQ ID NO: 2808) I037G01 26-35 50-66  99-114 SHYDILTRLNYWYFDL(SEQ ID NO: 2950) I037G03 26-35 50-66  99-119 DLGSFYDILTALRLENYGMDV(SEQ ID NO: 2963) I037G10 26-35 50-66  99-113 DYYDILTKLPYGMDV(SEQ ID NO: 2975) I042A07 26-35 50-66  99-117 VSPSYDILTGYYLPHAFDV(SEQ ID NO: 2849) I042A10 26-35 50-65  98-115 GPRYYDILTGYRYNWFDP(SEQ ID NO: 2801) I042B03 26-35 50-66  99-113 DIDDILTGYVLGMDV(SEQ ID NO: 2924) I042B12 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I042D01 26-35 50-66  99-109 QQWLPYDAFDI(SEQ ID NO: 2836) I042D03 26-35 50-68 101-113 AYYDILTGYFFDI(SEQ ID NO: 2873) I042D10 26-35 50-65  98-115 ERADYDILTGYYFYGMDV(SEQ ID NO: 2802) I042E10 26-37 50-69 102-120 ERPYYDILTGYTVTYGMDV(SEQ ID NO: 2798) I042E11 26-35 50-66  99-113 DEYDILTGLLQGMDV(SEQ ID NO: 2883) I042F08 26-37 50-67 100-115 GDYGILTGYPLHAFDI(SEQ ID NO: 2738) I042F12 26-35 50-66  99-113 DGYDILTGYYFGMDV(SEQ ID NO: 2976) I042G08 26-35 50-66  99-114 EHYDILTGYSLLGMDV(SEQ ID NO: 2907) I042G10 26-35 50-66  99-114 SHYDILTGLNYWYFDL(SEQ ID NO: 2166) I042H03 26-35 50-65  98-116 GSLYYDILTGYYIGNAFDI(SEQ ID NO: 2759) I043A03 26-35 50-66  99-117 DGYYDILTGGFYYYYGMDV(SEQ ID NO: 2899) I043B02 26-35 50-65  98-115 GGYYDILTGYLVYYGMDV(SEQ ID NO: 2744) I043B03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I043B06 26-35 50-66  99-116 DQQYDILTGYHIDYYMDV(SEQ ID NO: 2828) I043B07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I043B09 26-35 50-65  98-116 HVRDYDILTGYYRGHHFDY(SEQ ID NO: 2727) I043D11 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I043E05 26-35 50-66  99-116 TESNYDILTGYYWPSMDV(SEQ ID NO: 2940) I043F01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I043F04 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I043F12 26-35 50-66  99-116 TESNYDILTGYYWPSMDV(SEQ ID NO: 2940) I043H07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I044A11 26-35 50-68 101-117 APYDILTGYSDYYGMDV(SEQ ID NO: 2968) I044B11 26-35 50-66  99-112 DSDARLAALDAFDI(SEQ ID NO: 2978) I044C09 26-35 50-66  99-113 GQFGVLPNYYYHMDV(SEQ ID NO: 2943) I044C10 26-35 50-66  99-116 DIKRYNSNQPYYDYYMDV(SEQ ID NO: 2726) I044D03 26-35 50-66  99-117 DKQYYDILTGDPVEGGMDV(SEQ ID NO: 2889) I044D09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I044E07 26-35 50-66  99-110 AGSSLVTYGTDV(SEQ ID NO: 2825) I044E11 26-35 50-66  99-116 SDDYDILTGNYVGSLLDY(SEQ ID NO: 2758) I044F07 26-35 50-66  99-120 DGRLSYDILTGYYARDYYGMDV(SEQ ID NO: 2912) I044G02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I044G07 26-35 50-66  99-122 DQNHPIYDILTGYYVPTGPLELKN(SEQ ID NO: 2845) I044H01 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I050A01 26-35 50-66  99-114 DMGYDILTGYYGAFDI(SEQ ID NO: 2946) I050B12 26-35 50-66  99-114 DYYDVLTGFSLDGMDV(SEQ ID NO: 2829) I050006 26-35 50-65  98-113 DHYDVLTGSYLQAFDV(SEQ ID NO: 2728) I050008 26-37 52-67 100-114 GRYDFLTGYLRNFDY(SEQ ID NO: 2731) I050E01 26-35 50-66  99-113 GHYDILTGYYFGFDY(SEQ ID NO: 2886) I050E10 26-35 50-66  99-110 DMKVYYKYALDV(SEQ ID NO: 2823) I050H08 26-35 50-66  99-114 DLRYDILTGYHDAFDI(SEQ ID NO: 2890) I051A04 26-35 50-66  99-120 SSPPKWYDALTGHSSYHSAMDV(SEQ ID NO: 2159) I051A08 26-35 50-66  99-114 HRRARVVVPVPGAMDV(SEQ ID NO: 2930) I051Al2 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I051B08 26-36 51-67 100-115 RSMIVVTTAPYDAFDL(SEQ ID NO: 2785) I051C06 26-35 50-66  99-108 DTVRSGGMDV(SEQ ID NO: 2804) I051G12 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I055A05 26-35 50-66  99-106 SGPGWFDP (SEQ ID NO: 2870)I055A11 26-35 50-66  99-106 SGPGWFDP (SEQ ID NO: 2870) I061A03 26-3449-65  98-113 ELGSSIVGATTGALDM (SEQ ID NO: 2852) I061A04 26-35 50-66 99-114 GDYDILTGYPAECFQI (SEQ ID NO: 2854) I061A08 26-35 50-66  99-113DNYDILTGYSRRFDP (SEQ ID NO: 2942) I061A09 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I061A10 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I061B07 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I061B09 24-33 48-64  97-116EGGNYDILTGYYIGNGAFDI (SEQ ID NO: 2158) I061B12 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I061C12 26-35 50-66  99-111TYYDILTGYHFDY (SEQ ID NO: 2788) I061D01 26-35 50-68 101-110 GPGVIGNYDY(SEQ ID NO: 2749) I061D03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I061D04 26-35 50-66  99-113 AVLRYSAGLQGAFDI(SEQ ID NO: 2970) I061D07 26-35 50-66  99-114 VSGYNSGYFESYDMDV(SEQ ID NO: 2732) I061D09 26-35 50-66  99-114 LNLEKTVVRGFGYFDL(SEQ ID NO: 2952) I061D10 26-35 50-66  99-114 DHYDILTGLYYYGMDV(SEQ ID NO: 2760) I061E01 26-35 50-66  99-114 LNLEKTVVRGFGYFDL(SEQ ID NO: 2952) I061E05 26-35 50-66  99-115 GGELVWFGESDYYGMDV(SEQ ID NO: 2787) I061E09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I061E12 26-35 50-66  99-106 SQRLFIDS (SEQ ID NO: 2842)I061F01 26-35 50-66  99-119 DRYYDILTGYYIPGLDDAFDI (SEQ ID NO: 2887)I061F09 26-35 50-66  99-112 DSDARLAALDAFDI (SEQ ID NO: 2978) I061F1026-35 50-66  99-118 EESYYDILTGYYVHYYGMDV (SEQ ID NO: 2743) I061F11 26-3550-66  99-114 ATYDPLTGYYFDGFDI (SEQ ID NO: 2949) I061G01 26-35 50-66 99-114 ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I061G03 26-35 50-66  99-114AYYDILTGFLPYDMDL (SEQ ID NO: 2771) I061G09 26-35 50-66  99-117EVRNYDLLTRSYLAGPLDN (SEQ ID NO: 2751) I061G10 26-35 50-65  98-116EGSYDILTGYYVGVGRMDV (SEQ ID NO: 2171) I061G11 26-35 50-68 101-110RDILTGFYDS (SEQ ID NO: 2933) I061H05 26-37 50-67 100-115ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I064A05 26-35 50-68 101-115DFYDILTGYQHGMDV (SEQ ID NO: 2919) I064A11 26-35 50-66  99-111HSKEYNWNYALDY (SEQ ID NO: 2754) I064B01 26-35 50-66  99-111TRMDVLTRYYSDF (SEQ ID NO: 2750) I064B02 26-35 50-66  99-117AFEDYDILTGYYHHDAFDI (SEQ ID NO: 2911) I064B12 26-35 50-66  99-106PSYHYMDV (SEQ ID NO: 2740) I064C06 26-35 50-66  99-118VNADYDILTGYPRDYYGMDV (SEQ ID NO: 2819) I064D01 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I064D02 26-35 50-66  99-119EDATYYDILTGYYMGSYGMDV (SEQ ID NO: 2763) I064E01 26-35 50-66  99-116ETRKYTSSPPYNYYYMDV (SEQ ID NO: 2736) I064E02 26-35 50-66  99-113RDYDILTGYSRGFDP (SEQ ID NO: 2725) I064E03 26-35 50-66  99-117DGIYDILTTLVSYYNGMDV (SEQ ID NO: 2775) I064E07 26-35 50-65  98-113GERDILTGYYLDGMDV (SEQ ID NO: 2948) I064E08 26-35 50-66  99-113ERGSYSSGYSGAFDV (SEQ ID NO: 2898) I064F05 26-35 50-66  99-115ESGGYSYGSRDYYGMDV (SEQ ID NO: 2836) I064F08 26-35 50-66  99-118DRGVGYDILTGRTYYYGMDV (SEQ ID NO: 2900) I064G06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I065Al2 26-35 50-66  99-116DVSGHDILTGYSYRYFDV (SEQ ID NO: 2795) I065C04 26-35 50-66  99-112GQKNYYESSGYLEH (SEQ ID NO: 2916) I065C09 26-35 50-66  99-113GDYDILTGYYSHFDY (SEQ ID NO: 2908) I065E02 26-35 50-66  99-114AYDYDILTGYSYYFDY (SEQ ID NO: 2895) I065E04 26-35 50-66  99-108GMGDHYGMDV (SEQ ID NO: 2161) I065F03 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I065G06 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I065G07 26-35 50-66  99-115 GGNYDILTGYYIGAFDI(SEQ ID NO: 2824) I065G08 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I065H06 26-35 50-66  99-117 GYEYYDILTGYNELGAFDI(SEQ ID NO: 2851) I066A03 26-35 50-66  99-117 DGTYYDILTGYYNQYGMDV(SEQ ID NO: 2915) I066A08 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I066A09 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I066A10 26-35 50-66  99-115 DRGYDILTGYYYYGMDV(SEQ ID NO: 2876) I066A11 26-35 50-66  99-116 EVRDYDILTGYYISYMDV(SEQ ID NO: 2778) I066B02 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I066B08 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I066B10 26-35 50-66  99-115 GLYFEDTNYRHGDAFDI(SEQ ID NO: 2790) I066C02 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I066C11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I066C12 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I066D06 26-35 50-66  99-113 ENYDFLTGYYGAFDI(SEQ ID NO: 2772) I066D08 26-35 50-66  99-111 HSKEYNWNYALDY(SEQ ID NO: 2754) I066D11 26-35 50-66  99-117 ERSQFDFLTGVDRYHPMDV(SEQ ID NO: 2956) I066D12 26-35 50-66  99-112 EGAADYLNGQYFQH(SEQ ID NO: 2815) I066E06 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I066E12 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I066G05 26-35 50-66  99-115 GLYFEDTNYRHGDAFDI(SEQ ID NO: 2790) I066G08 26-35 50-66  99-114 VYYDILTGHPTYGMDV(SEQ ID NO: 2791) I066G10 26-35 50-68 101-117 GIYDILTGYHWDDAFDI(SEQ ID NO: 2872) I066G12 26-35 50-66  99-116 ESTYDILTGSYHDYGLDV(SEQ ID NO: 2822) I066H04 26-35 50-65  98-116 DRLHYDILTGHQTDDAFDI(SEQ ID NO: 2885) I067A07 26-35 50-66  99-117 VLTNYDILTGYYREDAFDM(SEQ ID NO: 2939) I067A11 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I067B08 26-35 50-66  99-122 DRGASNYDILTGYYAPAQGVAFDI(SEQ ID NO: 2969) I067C08 26-37 52-69 102-121 EGAHYDILTGHNYYHYGMDV(SEQ ID NO: 2747) I067C09 26-35 50-66  99-116 ETRKYTSSPPYNYYYMDV(SEQ ID NO: 2736) I067D07 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067E01 26-35 50-66  99-113 DQHDILTGVYYGMDV(SEQ ID NO: 2921) I067E06 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I067E07 26-35 50-67 100-123 DYPGSEYDILTGYLFGYYYYGMDV(SEQ ID NO: 2926) I067E11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I067G03 26-35 50-66  99-113 ARRVGVLGGKNAFEI(SEQ ID NO: 2765) I067G05 26-35 50-66  99-113 DQHDILTGGYYGMDV(SEQ ID NO: 2894) I067G12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I067H05 26-35 50-68 101-119 EGTYYDILTGYYPLGYFDY(SEQ ID NO: 2936) I067H06 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I068C09 26-35 50-66  99-110 GGSSQNFYGMDV(SEQ ID NO: 2884) I068G03 26-35 50-66  99-116 GTGYDILTGYYMGSAFDQ(SEQ ID NO: 2800) I068G04 26-35 50-66  99-115 GVVWVAYGDVGIYGFDV(SEQ ID NO: 2937) I068G07 26-35 50-66  99-113 HDYYIMTAAHYYYDS(SEQ ID NO: 2909) I068G08 26-35 50-66  99-116 GIGYDLLTGYFTGSPLDY(SEQ ID NO: 2846) I070F07 26-35 50-66  99-113 DFYDILTGYHDAFDI(SEQ ID NO: 2910) I070G05 26-35 50-68 101-113 DVDDILTGYSWDY(SEQ ID NO: 2867) I070H02 26-35 50-66  99-114 MEYDILTGYYGGYFDY(SEQ ID NO: 2179) I071A01 26-35 50-66  99-114 AAYDPLTGYSFDGFDI(SEQ ID NO: 2783) I071A03 26-35 50-66  99-116 DMHYDILTGYYTGLAFDM(SEQ ID NO: 2917) I071B08 27-36 51-67 100-115 GGYDILTQYPAEFFHP(SEQ ID NO: 2764) I071E01 26-35 50-66  99-111 DFGVIGDYRPFDY(SEQ ID NO: 2777) I071F11 26-35 50-66  99-108 SSNPVYGLDV(SEQ ID NO: 2957) I071G11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I071H08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I074A02 26-35 50-66  99-114 DDRDILTNYYLEYFQH(SEQ ID NO: 2868) I074A08 26-35 50-66  99-120 SSPPKWYDALTGDSSYHSAMDV(SEQ ID NO: 2165) I074D10 26-35 50-66  99-117 DKTLGDQLVEAYYYDGMDV(SEQ ID NO: 2776) I074E01 26-35 50-66  99-117 LGRTSRDLLTGYHFYNMDV(SEQ ID NO: 2944) I074E02 26-35 50-66  99-113 DDYDILTGSLYYFDS(SEQ ID NO: 2803) I074E08 26-35 50-66  99-116 GTGYDILTGYYMGSAFDQ(SEQ ID NO: 2800) I074F12 26-35 50-66  99-113 DRADILTGYNDAFDI(SEQ ID NO: 2739) I074H06 26-35 50-66  99-112 RYGDPFYYYYYMNV(SEQ ID NO: 2755) I074H07 26-35 50-66  99-116 GTGYDILTGYYMGSAFDQ(SEQ ID NO: 2800) I074H08 26-35 50-66  99-115 VSNDILTGWGGYNWFDP(SEQ ID NO: 2955) I075A07 26-35 50-66  99-116 GTGYDILTGYYMGSAFDQ(SEQ ID NO: 2800) I075B01 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I075B04 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175) I075B06 26-3449-65  98-113 ELGLSIVGATTGALDM (SEQ ID NO: 2174) I075B08 26-35 50-66 99-116 GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I075B09 26-35 50-66  99-114TYYDILTGYYAEYFQH (SEQ ID NO: 2932) I075B12 26-35 50-66  99-113DDYDILTGYYWVPAV (SEQ ID NO: 2812) I075C01 26-35 50-66  99-120GREDTDKVKPWDRYFHYYYMDV (SEQ ID NO: 2835) I075C05 26-35 50-66  99-106DQGRYLDL (SEQ ID NO: 2175) I075D05 26-35 50-66  99-116GTGYDILTGYYMGSVFDP (SEQ ID NO: 2897) I075D07 26-35 50-66  99-114SYYDILTGYYHTPLDY (SEQ ID NO: 2853) I075D08 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I075E01 26-35 50-66  99-116GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I075E03 28-37 52-68 101-121GGGYDILTGYSYPYLYYGLDV (SEQ ID NO: 2865) I075E04 26-35 50-66  99-116GRGYDVLTGYFTGSPLDY (SEQ ID NO: 2881) I075E05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I075E10 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I075E11 26-35 50-66  99-106 SGPGWFDP(SEQ ID NO: 2870) I075E12 26-35 50-66  99-115 TDRFGAKDVTARWGMDV(SEQ ID NO: 2979) I075F02 26-35 50-66  99-117 EQGYDILTGYYPEGGWFDP(SEQ ID NO: 2834) I075F04 26-37 52-67 100-114 AGYDLLTGYPFYFDS(SEQ ID NO: 2757) I075F06 26-35 50-66  99-117 GRNYYDFLTGYNFNLGLDY(SEQ ID NO: 2830) I075F07 26-35 50-66  99-113 ENYDSLTGYYNYFDY(SEQ ID NO: 2971) I075F08 26-35 50-66  99-106 DQRKAQDI (SEQ ID NO: 2779)I075F09 26-35 50-66  99-118 LKAPYYDLLTGYHLPKWFDT (SEQ ID NO: 2953)I075F10 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175) I075F11 26-3550-66  99-106 DQGRYLDL (SEQ ID NO: 2175) I075G05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I075G07 26-35 50-66  99-113GRYYDMLTRGGYFDY (SEQ ID NO: 2858) I075G08 26-35 50-66  99-113RQYDILTGYYGGFDY (SEQ ID NO: 2958) I075G11 26-35 50-66  99-114TDYDILTGYPMGYFDP (SEQ ID NO: 2173) I075G12 26-35 50-66  99-106 DQGRYLDL(SEQ ID NO: 2175) I075H02 26-35 50-66  99-116 GTGYDILTGYYMGSAFDQ(SEQ ID NO: 2800) I075H03 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I075H06 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175) I075H08 26-3550-66  99-116 GSGYDLLTGYFTGSPLDY (SEQ ID NO: 2766) I076A01 26-35 50-66 99-115 DRRRDDLTGYLYDAFDS (SEQ ID NO: 2878) I076A03 26-35 50-68 101-108GYDTAMQY (SEQ ID NO: 2951) I076A06 26-35 50-66  99-106 DQGRYLDL(SEQ ID NO: 2175) I076A07 26-35 50-66  99-112 DRRDILTGSNFGQD(SEQ ID NO: 2913) I076A08 26-35 50-66  99-115 MGHYDILTGYRHYGMDV(SEQ ID NO: 2831) I076B01 26-35 50-66  99-116 GSGYDLLTGYFTGSPLDY(SEQ ID NO: 2766) I076B03 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I076B07 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175) I076B08 26-3550-66  99-114 PYYDPLTAYTFQYFGN (SEQ ID NO: 2806) I076C04 26-34 49-65 98-113 ELGLSIVGATTGALDM (SEQ ID NO: 2174) I076C10 26-35 50-66  99-113GRYYDMLTRGGYFDY (SEQ ID NO: 2858) I076D01 26-35 50-66  99-114LDYDILTGYYPSGFDY (SEQ ID NO: 2799) I076D08 26-37 52-67 100-113RFYDLLTGYSAFDS (SEQ ID NO: 2756) I076D11 26-35 50-66  99-116GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I076D12 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I076E04 26-35 50-66  99-116GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I076E07 26-35 50-66  99-113EYYDVLTGLFYYMDV (SEQ ID NO: 2841) I076E09 26-35 50-66  99-114DDRDILTNYYLEYFQH (SEQ ID NO: 2868) I076E11 26-35 50-66  99-116GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I076F01 26-35 50-66  99-116GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I076F03 26-36 51-66  99-113GDYDVLTGYLRKLDY (SEQ ID NO: 2742) I076F04 26-35 50-66  99-106 DQGRYLDL(SEQ ID NO: 2175) I076F08 26-36 51-66  99-113 VHYDILTGYLWAFDI(SEQ ID NO: 2730) I076F10 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I076G09 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I076G10 26-35 50-66  99-113 GRYYDMLTRGGYFDY (SEQ ID NO: 2858) I076G1126-35 50-66  99-116 GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I076G12 26-3550-66  99-119 NGYYDILTGYYLWDYYYGMDV (SEQ ID NO: 2769) I076H02 26-3550-66  99-113 ENYDSLTGYYNYFDY (SEQ ID NO: 2971) I076H04 26-35 50-66 99-114 THYDILTGYYSHPLDY (SEQ ID NO: 2863) I076H05 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I076H06 26-35 50-66  99-113VPYDILTGYWGAFDV (SEQ ID NO: 2827) I076H09 26-35 50-66  99-116GSGYDLLTGYFTGSPLDY (SEQ ID NO: 2766) I076H10 26-35 50-66  99-116GSGYDLLTGYFTGSPLDY (SEQ ID NO: 2766) I077D06 26-35 50-66  99-113VYYDILTGYNLFFDY (SEQ ID NO: 2177) I078B04 26-35 50-66  99-113VYYDILTGYNLFFDY (SEQ ID NO: 2177) I078E10 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I002A01-K 26-35 50-66  99-114ELGLSIVGATTGALDM (SEQ ID NO: 2174) I002A01-R 26-35 50-66  99-114ELGLSIVGATTGALDM (SEQ ID NO: 2174) I026C04-K 26-35 50-66  99-114ELGLSIVGATTGALDM (SEQ ID NO: 2174) I026C04-R 26-35 50-66  99-114ELGLSIVGATTGALDM (SEQ ID NO: 2174) I067B10 26-35 50-66  99-122DRGAPNYDILTGYYAPAQGVARDI (SEQ ID NO: 2176) I068C06 26-35 50-66  99-106DQGRYLDL (SEQ ID NO: 2175) I075F12 26-35 50-66  99-106 DQGRYLDL(SEQ ID NO: 2175) I003C06 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I025B06 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I025B09 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I026C04 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I027B12 26-34 49-65  99-114 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I030A10 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I064C04 26-35 50-66  99-120 DGRLSYDILTGYYARDYYGMDD(SEQ ID NO: 2188) I064C07 26-35 50-66  99-107 SEGTIFGVD(SEQ ID NO: 2178) I065D04 26-36 51-66  99-117 GKGYYDILTGYYRDNWFDP(SEQ ID NO: 2181) I065D08 26-35 50-66  99-120 TPSSVYDLLTGYYHYFYSYMDV(SEQ ID NO: 2189) I065F08 26-35 50-66  99-108 EKSAAGYFDY(SEQ ID NO: 2190) I067F05 26-35 50-66  99-113 ENYDSLTGYYGAFDI(SEQ ID NO: 2185) I068B04 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I068B08 26-34 49-65  98-113 KLGLSIVGATTGALDM (SEQ ID NO: 2186) I068C0826-35 50-66  99-115 EGMNDFINSHHYYTMDA (SEQ ID NO: 2182) I068F03 26-3550-66  99-112 AGNEYGHTERPADY (SEQ ID NO: 2180) I069B07 26-35 50-66 99-114 MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I071B03 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I072B09 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I073F04 26-35 50-66  99-109SLATRPLGMDV (SEQ ID NO: 2184) I074B12 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I075A02 26-34 49-65  98-113ELGLSIVGATTGALDM (SEQ ID NO: 2174) I075G01 26-35 50-66  99-113DHFDTLTGYFRRLDS (SEQ ID NO: 2187) I078D02 26-35 50-66  99-113VYYDILTGYNLFFDY (SEQ ID NO: 2177) I078D08 26-35 50-66  99-117DAQSYYDILTGYQSYAFDI (SEQ ID NO: 2183) I078H08 26-35 50-66  99-113VYYDILTGYNLFFDY (SEQ ID NO: 2177) I064A03 26-35 50-66  99-123GPSTTYYDILTGYYTPYYYYYYMDV (SEQ ID NO: 3014) I064B03 26-37 52-67 100-118HVRDYDILTGYYRGHYFDY (SEQ ID NO: 2167) I064B05 26-35 50-66  99-113ERGVVTAYGGDSFDL (SEQ ID NO: 2985) I064B11 26-35 50-66  99-111DRGPGLLSSFFES (SEQ ID NO: 3033) I064C02 26-35 50-66  99-119DEYYDILTGYQAPYYYYGMDV (SEQ ID NO: 3068) I064C03 26-35 50-66  99-113ERGVVTAYGGDSFDL (SEQ ID NO: 2985) I064C11 26-35 50-66  98-116DVTYHDILTGYAGHEAFDI (SEQ ID NO: 3055) I064C12 26-37 52-69 102-121ESGRYDILTGYYSGGGGMDV (SEQ ID NO: 3012) I064D03 26-35 50-66  99-119DGANYDILTGYYTTTVYGMDV (SEQ ID NO: 3072) I064D04 26-35 50-66  99-114RSYDILTGYYTYGMDV (SEQ ID NO: 3090) I064D06 26-35 50-66  99-107 EGSSGYLVG(SEQ ID NO: 2981) I064E05 26-37 52-67 100-119 KQRGDYDILTGYQLGYAFDI(SEQ ID NO: 2808) I064E06 26-35 50-66  99-118 ERPGYDILTGYPSSIYGMDV(SEQ ID NO: 3053) I064F07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I064F09 26-35 50-66  99-120 DTLGYDILTGYPPPYYYYYDMDV(SEQ ID NO: 2988) I064F10 22-31 46-62  95-116 DTLGYDILTGYPPPYYYYDMDV(SEQ ID NO: 2988) I064F11 26-35 50-65  98-115 GRHYYDILTGYYNEAFDI(SEQ ID NO: 3031) I064G01 26-35 50-66  99-113 NYYDVLTQSYYGMDV(SEQ ID NO: 3077) I064G04 26-35 50-66  99-106 DNSGTYGY (SEQ ID NO: 3084)I064G08 26-35 50-66  99-111 GGVTAGRSVYFDS (SEQ ID NO: 2990) I064G1026-35 50-66  99-113 SPNGDYSGYAWGLEY (SEQ ID NO: 3085) I064G11 26-3550-65  98-111 YRDGSGYYPVSFSY (SEQ ID NO: 3064) I064G12 26-35 50-65 98-112 VNYDILTGLGYYFDY (SEQ ID NO: 3049) I064H03 26-37 50-67 100-116SYYDILTGRPYTDAFDI (SEQ ID NO: 2989) I064H04 26-35 50-66  99-115PLGITAVRGAKTDAFGI (SEQ ID NO: 2929) I064H06 26-35 50-66  99-122DRGASNYDILTGYYAPAGQVAFDI (SEQ ID NO: 2969) I065A02 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I065A04 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I065A06 26-35 50-66  99-114ATYDPLTGYSFDGFDI (SEQ ID NO: 2153) I065A07 26-35 50-66  99-117DGGGYDILTGYQYYYGMDV (SEQ ID NO: 2987) I065B01 26-35 50-65  98-118WATYYDTLTGYRLKDHAGFDI (SEQ ID NO: 3017) I065B05 26-35 50-66  99-115SPGDDILTGYYKYYFDY (SEQ ID NO: 3032) I065B09 26-35 50-66  99-119DAGESYDILTGYYVIEGYMDV (SEQ ID NO: 2986) I065B12 26-35 50-66  99-112EGAADYLNGQYFQH (SEQ ID NO: 2815) I065C02 26-35 50-66  99-109 EGSWSGLDLDY(SEQ ID NO: 3007) I065C06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I065C08 26-35 50-66  99-114 VSGYNSGYFESYDMDV(SEQ ID NO: 2732) I065C10 26-35 50-66  99-110 QGGQYDSPPLDV(SEQ ID NO: 3002) I065D01 26-35 50-66  99-115 DRDYDILTDYSNYGMDV(SEQ ID NO: 3074) I065D03 26-35 50-66  99-115 APLYDILTGYYIGGNDY(SEQ ID NO: 3028) I065D05 26-35 50-66  99-116 DKDYDILTGYWRDELLDY(SEQ ID NO: 3040) I065D06 26-35 50-66  99-115 DPNYDILTGYYYYAMDV(SEQ ID NO: 3062) I065E01 26-35 50-66  99-112 EFDQLLARGHGMDV(SEQ ID NO: 3027) I065E05 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I065E06 26-35 50-66  99-119 ARGSYDILTGYYRPGDGYFDY(SEQ ID NO: 3043) I065E08 26-35 50-66  99-115 GLYFEDTNYRHGDAFDI(SEQ ID NO: 2790) I065E09 26-35 50-65  98-118 RESYYDILTGYSPRSKYGMDV(SEQ ID NO: 3021) I065E12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I065F04 26-35 50-66  99-113 ERGVVTAYGGDSFDL(SEQ ID NO: 2985) I065F05 26-35 50-65  98-113 RYSDALTGYSLGAFDV(SEQ ID NO: 3018) I065F07 26-38 53-69 102-118 GAYYDILTGYYPYGMDV(SEQ ID NO: 2860) I065F09 26-35 50-66  99-116 DYPIDVLTGRRTKNWFDP(SEQ ID NO: 3013) I065F12 26-35 50-66  99-114 DQVDRLLMQYNYYMDA(SEQ ID NO: 3047) I065G01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I065G09 26-35 50-68 101-116 DAYYDILTGWVYGMDV(SEQ ID NO: 3030) I065G10 26-36 51-66  99-113 FRYDILTGYYYDMDV(SEQ ID NO: 2983) I065H05 26-35 50-66  99-113 EYYDILTGYSGAFDI(SEQ ID NO: 2984) I065H07 26-35 50-66  99-111 TRMDVLTRYYSDF(SEQ ID NO: 2750) I066A05 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I066A06 26-35 50-66  99-112 EGAADYLNGQYFQH(SEQ ID NO: 2815) I066Al2 26-35 50-66  99-115 DTRVIGIQLWERGAFDM(SEQ ID NO: 3080) I066B05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I066B11 26-35 50-66  99-115 PLGITAVRGAKTDAFGI(SEQ ID NO: 2929) I066C06 26-35 50-65  99-117 GRRYYDILTGYSLGRGEMDV(SEQ ID NO: 3009) I066C10 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I066D02 26-35 50-66  99-110 AGTSLMNYGTDV(SEQ ID NO: 3048) I066D07 26-35 50-66  99-114 GPYDVLTGYLSGNFDY(SEQ ID NO: 2992) I066E01 26-35 50-66  99-110 QGGQYDSPPFDV(SEQ ID NO: 3001) I066E03 26-35 50-66  99-122 GEKARYYDILTGYYSAWGGYYMDV(SEQ ID NO: 3045) I066E04 26-35 50-66  99-114 LNLEKTVIRGFGYFDL(SEQ ID NO: 3081) I066E05 26-35 50-66  99-115 VGGYDILTGYYLRGMDV(SEQ ID NO: 2997) I066E07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I066E09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I066F01 26-35 50-66  99-114 SPYDTLTGYVYNGVDV(SEQ ID NO: 3058) I066F03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I066F04 26-35 50-66  99-114 VAAAGARTLGYFGMDV(SEQ ID NO: 3071) I066F07 26-35 50-66  99-116 DVSGHDILTGYSYRYFDV(SEQ ID NO: 2795) I066F08 26-35 50-66  99-117 SPMYYDRLTGFYPSGYFDS(SEQ ID NO: 3036) I066F11 26-35 50-66  99-115 GAYYDILTGYYPYGMDV(SEQ ID NO: 2860) I066F12 26-35 50-66  99-114 GPSSAGTTIGLGSFDP(SEQ ID NO: 3005) I066G06 26-35 50-66  99-116 ETRKYTSSPPYNYYYMDV(SEQ ID NO: 2736) I066G07 26-30 45-61  94-106 DQFSVGGRHAFDL(SEQ ID NO: 3054) I066H02 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I067A02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I067A03 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067A06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I067A08 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067A10 26-35 50-66  99-113 ERGVVTAYGGDSFDL(SEQ ID NO: 2985) I067B03 26-35 50-66  99-115 PLGITAVRGAKTDAFGI(SEQ ID NO: 2929) I067B04 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067C03 26-35 50-66  99-106 DWGHWFDP (SEQ ID NO: 2982)I067C05 26-35 50-66  99-110 SGSSLMTYGTDV (SEQ ID NO: 3015) I067C07 26-3550-66  99-114 EPYDILTGYYGSYFDY (SEQ ID NO: 3041) I067C10 26-35 50-66 99-110 AGSSLMTYGTDV (SEQ ID NO: 2773) I067C12 26-35 50-66  99-115TYYDILTGYSGGGAFDY (SEQ ID NO: 3024) I067D01 26-35 50-66  99-109GSRVRGVTPDL (SEQ ID NO: 3020) I067D03 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067D05 26-35 50-66  99-119 ECSGSSCPARQPPYYQYYMDV(SEQ ID NO: 2993) I067D06 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067D09 26-35 50-66  99-115 GATTDILTGYYPYGMDV(SEQ ID NO: 2860) I067D12 26-35 50-66  99-110 QGGQYDSPPLDV(SEQ ID NO: 3002) I067E02 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067E04 26-35 50-66  99-115 GAYYDILTGYYPYGMDV(SEQ ID NO: 2860) I067E05 26-35 50-66  99-117 DYRNYDILTGHPYYYGMDV(SEQ ID NO: 2996) I067F01 26-35 50-66  99-114 QHYDILTGYSQEPFDI(SEQ ID NO: 3022) I067F03 26-35 50-66  99-117 DQTYYDILTGHYYYYGMDV(SEQ ID NO: 3087) I067F04 26-35 50-66  99-112 EGAADYLNGQYFQH(SEQ ID NO: 2815) I067F08 26-35 50-66  99-113 LGYYDILTGYRSDDY(SEQ ID NO: 3029) I067F10 26-35 50-66  99-110 AGSSLMAYGTDV(SEQ ID NO: 3016) I067F11 26-35 50-66  99-113 ENYDFLTGYYGAFDI(SEQ ID NO: 2772) I067G01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I067G09 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I067H07 26-35 50-66  99-117 GGLYDILTGRPATDDAFDI(SEQ ID NO: 3035) I068A07 26-35 50-66  99-115 TDRFGAKDVARWGMDV(SEQ ID NO: 2979) I068E05 26-35 50-66  99-120 GREDTDKVKPWDRYYHYYYMDV(SEQ ID NO: 2809) I068E08 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I068E11 26-34 49-65  98-113 ELGLSIVGATTGALDM (SEQ ID NO: 2174) I068F0426-35 50-66  99-114 ELGHREGGYWYSPYNV (SEQ ID NO: 2838) I068G05 26-3550-66  98-108 KNMGASAAADF (SEQ ID NO: 3042) I068G06 26-35 50-66  99-112RYGDPFYYYYYMNV (SEQ ID NO: 2755) I068G11 26-35 50-66  99-119ESGSHYDLLTGLLVAANGFDV (SEQ ID NO: 3044) I069A09 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I069A10 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I069B06 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I069B09 26-35 50-66  99-112PYYDILTGYFAFDI (SEQ ID NO: 3026) I069B12 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I069C06 26-35 50-66  99-116VLPHYDILTGYSQNWFDP (SEQ ID NO: 3000) I069C09 26-35 50-66  99-116VLPHYDILTGYSQNWFDP (SEQ ID NO: 3000) I069D03 26-35 50-66  99-115DGYYDILTGYSYYGMDV (SEQ ID NO: 2135) I069E09 26-35 50-66  99-115DGYYDILTGYSYYGMDV (SEQ ID NO: 2135) I069E11 26-35 50-66  99-113VYYDILTGYNLFFDY (SEQ ID NO: 2177) I069F05 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I069F07 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I069F12 26-35 50-66  99-113GYYDILTGYYDAFDI (SEQ ID NO: 3051) I069G06 26-35 50-66  99-115DGYYDILTGYSGYYMDV (SEQ ID NO: 3059) I069G08 26-35 50-66  99-118DRLEYYDILTGYYYYYGMDV (SEQ ID NO: 3039) I069G11 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I070A03 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I070A09 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I070B01 26-35 50-66  99-117SQSDYDILTGYYYYYGMDV (SEQ ID NO: 3038) I070B05 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I070D03 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I070D04 26-35 50-66  99-114MEYDILTSYYGGYFDY (SEQ ID NO: 3034) I070E01 26-35 50-66  99-117SQSDYDILTGYYYYYGMDV (SEQ ID NO: 3038) I070F01 26-35 50-66  99-117SQSNYDILTGYYYYYGMDV (SEQ ID NO: 3067) I070G10 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I071A06 26-35 50-66  99-108GMGDHYGMDV (SEQ ID NO: 2161) I071B02 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I071D02 26-35 50-66  99-110 AGTSLMNYGTDV(SEQ ID NO: 3048) I071D08 26-37 52-66  99-119 VPYYYDTSGGYLGEYYYGMDV(SEQ ID NO: 3010) I071F01 26-35 50-66  99-110 AGTSLMNYGTDV(SEQ ID NO: 3048) I071G09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072A01 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I072A09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072B02 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I072B10 26-35 50-66  99-110 AGSSLMTYGTDV(SEQ ID NO: 2773) I072B11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072B12 26-35 50-66  99-113 ENYDYLTGYYGAFDI(SEQ ID NO: 2995) I072C05 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I072C10 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072D01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072D05 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I072E01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072E04 26-35 50-66  99-117 EGSYDILTGYYVGVGRMDV(SEQ ID NO: 2171) I072E05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072E06 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I072F03 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I072F07 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072F11 26-35 50-66  99-113 DEYDILTGLLQGMDV(SEQ ID NO: 2883) I072G03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072G04 26-35 50-68 101-110 RDILTGFYDS(SEQ ID NO: 2933) I072G05 26-35 50-66  99-110 GYRNDWYGAFEI(SEQ ID NO: 3079) I072G09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072H03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I072H07 26-35 50-66  99-110 AGTSLMNYGMDV(SEQ ID NO: 3070) I073A02 26-35 50-66  99-114 GPYDILTGYYRDAFDI(SEQ ID NO: 2998) I073A03 26-35 50-66  99-115 THYDILTGYYTADAFDI(SEQ ID NO: 3019) I073A04 26-35 50-66  99-121 VQMDSEYYDLLTGINVGPYYFDY(SEQ ID NO: 2132) I073A05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073A06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073A09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073A10 26-35 50-66  99-119 GDFGDYDILTGYYPVYYGMDV(SEQ ID NO: 3082) I073A11 26-35 50-66  99-114 SYYDILTGYYPFGMDV(SEQ ID NO: 3004) I073B02 26-35 50-66  99-117 DLWYYDILTGYYLDDAFDI(SEQ ID NO: 2999) I073B05 26-35 50-66  99-117 DLWYYDILTGYYLDDAFDI(SEQ ID NO: 2999) I073B06 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I073B07 26-35 50-66  99-111 TRMDVLTRYYSDF(SEQ ID NO: 2750) I073B08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073B11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073C01 26-35 50-66  99-114 GYHDTLTSYNYNWFDP(SEQ ID NO: 3006) I073C02 26-35 50-66  99-121 AQMDSEYYDLLTGINVGPYYFDY(SEQ ID NO: 3076) I073C04 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073C07 26-35 50-66  99-107 GMGDHYMDV(SEQ ID NO: 3008) I073C08 26-35 50-66  99-115 EMGYDILTGYYLNYMDV(SEQ ID NO: 2862) I073C09 26-35 50-66  99-114 QHYDILTGYSQEPFDI(SEQ ID NO: 3022) I073C11 26-35 50-68 101-119 FNPTYDILTGYYIGGYFQH(SEQ ID NO: 2155) I073C12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073D01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073D03 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I073D06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073D08 26-35 50-66  99-117 EVRNYDLLTRSYLAGPLDN(SEQ ID NO: 2751) I073D10 26-35 50-68 101-113 QYYDILTGYELDI(SEQ ID NO: 3073) I073D11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073E01 26-37 52-69 102-121 EGAHYDILTGHNYYHYGMDV(SEQ ID NO: 2747) I073E02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073E03 26-35 50-66  99-114 ATYDPLTGYSLDGFDI(SEQ ID NO: 3003) I073E05 26-35 50-66  99-114 QHYDILTGYSQEPFDI(SEQ ID NO: 3022) I073E06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073E08 26-35 50-66  99-113 ENYDFLTGYYGAFDI(SEQ ID NO: 2772) I073F01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073F02 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073F03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073F05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073F07 26-35 50-66  99-114 GEYDILTGYPYWYFDL(SEQ ID NO: 3023) I073F09 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073F11 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073F12 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073G03 26-35 50-66  99-116 DGSYDILTGYYIDNYMDV(SEQ ID NO: 2154) I073G04 26-35 50-65  98-116 GEGGYDILTGYLRGYGMDV(SEQ ID NO: 3037) I073G05 26-35 50-66  99-108 GMGDHYGMDV(SEQ ID NO: 2161) I073G06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073G07 26-35 50-66  99-115 GSYYDILTGISSLGMDV(SEQ ID NO: 3063) I073G08 26-35 50-66  99-112 SRDLLLFPHYGMDV(SEQ ID NO: 2133) I073G09 26-35 50-66  99-118 DRGHYDILTGYYIEPSGFDY(SEQ ID NO: 3061) I073G10 26-35 50-66  99-108 GPGVIGNYDY(SEQ ID NO: 2749) I073G12 26-35 50-68 101-115 GGMIRAREDYYYMDV(SEQ ID NO: 3083) I073H01 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073H03 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073H05 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073H06 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I073H07 26-35 50-66  99-111 TYYDILTGYYFDY(SEQ ID NO: 3056) I073H08 26-35 50-66  99-114 ATYDPLTGYSFDGFDI(SEQ ID NO: 2153) I074A05 26-35 50-66  99-116 LPPYDMLTGYYVGGGMDV(SEQ ID NO: 3050) I074A06 26-35 50-66  99-116 AKPYTDFSRGSDADAFDV(SEQ ID NO: 3065) I074B03 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I074B11 26-35 50-66  99-112 RYGDPFYYYYYMNV (SEQ ID NO: 2755) I074C0726-34 49-65  98-113 ELGLSIVGATTGALDM (SEQ ID NO: 2174) I074D03 26-3550-66  99-114 GGYDILTQYPAEFFHP (SEQ ID NO: 2764) I074D04 26-35 50-66 99-106 DQGRYLDL (SEQ ID NO: 2175) I074D05 26-35 50-66  99-116DRYYDILTKGDYYYGMDV (SEQ ID NO: 3060) I074D07 26-35 50-66  99-123VQGETYYDILTGYWGPKRDLYGMDV (SEQ ID NO: 3069) I074D08 26-34 49-65  98-113ELGLSIVVATTGALDM (SEQ ID NO: 2980) I074D11 26-35 50-66  99-111ESEGGDYTNPFGY (SEQ ID NO: 2991) I074E05 26-35 50-66  99-106 DQGRYLDL(SEQ ID NO: 2175) I074E07 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I074E09 26-35 50-68 101-119 DPGNYDILTGYYYYYGMDV(SEQ ID NO: 2935) I074E11 26-35 50-66  99-110 VRLPHHHYFMAV(SEQ ID NO: 3075) I074H05 26-35 50-66  99-115 ESSITVNPPYYFYGMDV(SEQ ID NO: 3025) I075A03 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I075A10 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175) I075B07 26-3550-66  99-116 DPEGDYQPLSSNYNWLDP (SEQ ID NO: 3011) I075D11 26-36 51-66 99-106 GKEGYNDN (SEQ ID NO: 3089) I075D12 26-35 50-66  99-116GSGYDLLTGYFTGSPLDY (SEQ ID NO: 2766) I075G02 26-35 50-66  99-116SPEGDYQPLSSNYNWLDP (SEQ ID NO: 3011) I075G09 26-35 50-66  99-115MGHYDILTGYRHYGMDV (SEQ ID NO: 2831) I075G10 26-35 50-66  99-111GNYDILTGYPHDL (SEQ ID NO: 3086) I075H05 26-35 50-66  99-114SYYDILTGYYHTPLDY (SEQ ID NO: 2853) I075H07 26-35 50-66  99-116GSGYDLLTGYFTGSPLDY (SEQ ID NO: 2766) I076A11 26-35 50-66  99-114DDRDILTNYYLEYFQH (SEQ ID NO: 2868) I076Al2 26-35 50-66  99-116GSGYDVLTGYFTGSPLDY (SEQ ID NO: 3057) I076B06 26-35 50-66  99-113GRYDILTGYFTSFDY (SEQ ID NO: 3066) I076B10 26-35 50-66  99-114DDRDILTNYYLEYFQH (SEQ ID NO: 2868) I076B12 26-35 50-66  99-116GTGYDILTGYYMGSAFDQ (SEQ ID NO: 2800) I076C06 26-35 50-66  99-115MGHYDILTGYRHYGMDV (SEQ ID NO: 2831) I076C11 26-35 50-66  99-106 DQGRYLDL(SEQ ID NO: 2175) I076D06 26-34 49-65  98-113 ELGLSIVGATTGALDM(SEQ ID NO: 2174) I076E05 26-35 50-66  99-116 GTGYDILTGYYMGSAFDQ(SEQ ID NO: 2800) I076E08 26-35 50-66  99-106 DQGRYLDL (SEQ ID NO: 2175)I076F06 26-36 51-66  99-106 RDVQGAPY (SEQ ID NO: 3088) I076G01 26-3550-66  99-116 VEGVYDILTGYSFDAFDI (SEQ ID NO: 3078) I076H01 26-35 50-66 99-117 EQGYDILTGYYPEGGWFDP (SEQ ID NO: 2834) I076H03 26-34 49-65 99-113 ELGLSIVGATTGALDM (SEQ ID NO: 2174) I077B05 26-37 52-69 102-120DKSYYDILTGYYYYYGMDV (SEQ ID NO: 3052) I077C10 26-35 50-66  99-114MEYDILTGYYYGGYFDY (SEQ ID NO: 2179) I077D01 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I077D04 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I077D11 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I077D12 26-35 50-66  99-113EKYDILTGYYDAFDI (SEQ ID NO: 3046) I077E01 26-35 50-66  99-115EMGYDILTGYYLNYMDV (SEQ ID NO: 2862) I077E03 26-35 50-66  99-115EMGYDILTGYYLNYMDV (SEQ ID NO: 2862) I077E08 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I077F05 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I077G06 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I077H02 26-35 50-66  99-114MEYDILTGYYGGYFDY (SEQ ID NO: 2179) I078B05 26-35 50-66  99-116ESHYDILTGYYSNPSFDI (SEQ ID NO: 2994) I079E02 26-35 50-66  99-110DSGSYYYDAFDI (SEQ ID NO: 2194) I079F11 26-35 50-66  99-105 TGSGFDY(SEQ ID NO: 2192) I082G02 26-35 50-66  99-109 DGYRTNDALDI(SEQ ID NO: 2191) I082H08 26-35 50-66  99-104 DWDMDV (SEQ ID NO: 2193)I099D03 26-35 50-66  99-109 DNGGGTIGFDY (SEQ ID NO: 2195) I079B05 26-3550-66  99-103 FVLDY (SEQ ID NO: 2210) I079B12 26-35 50-66  99-107WTSSGAFDI (SEQ ID NO: 2205) I079C01 26-35 50-66  99-104 DWDMDV(SEQ ID NO: 2193) I079F06 26-35 50-66  99-107 DNLHAAFDI(SEQ ID NO: 2202) I079F08 26-35 50-66  99-111 YYYHSSGSDAFDI(SEQ ID NO: 2206) I080A03 26-35 50-66  99-111 VGIKAAAVDNFEY(SEQ ID NO: 2197) I080A08 26-35 50-66  99-108 VHSTGYAFEN(SEQ ID NO: 2200) I080B01 26-35 50-66  99-115 EYSGYHYVEGGSYAMDV(SEQ ID NO: 2201) I080D03 26-35 50-66  99-111 VGIKAAAVDNFEY(SEQ ID NO: 2197) I080E05 26-35 50-66  99-114 EGGGDAYDVAPYYFDY(SEQ ID NO: 2204) I080G07 26-35 50-66  99-109 EGPGYYYGMDV(SEQ ID NO: 2209) I080G09 26-35 50-66  99-109 DNGGGTIGFDY(SEQ ID NO: 2195) I082A05 26-35 50-66  99-104 DLDFDY (SEQ ID NO: 2208)I082B08 26-35 50-66  99-110 DLGIAGTIYFDY (SEQ ID NO: 2207) I082C03 26-3550-66  99-111 DASRDIVVLPLAI (SEQ ID NO: 2198) I082D07 26-35 50-66 99-107 WTSSGAFDI (SEQ ID NO: 2205) I082G01 26-35 50-66  99-111DRGSGWPNWYFDL (SEQ ID NO: 2212) I083B12 26-35 50-66  99-110 ESGAGGYYYDDY(SEQ ID NO: 2196) I083G03 26-35 50-66  99-111 VGIKAAAVDNFEY(SEQ ID NO: 2197) I084A01 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I084B02 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203) I084C04 25-34 49-65 98-104 NLWGLDY (SEQ ID NO: 2199) I084C11 26-35 50-66  99-107 GNAWGAFDI(SEQ ID NO: 2211) I079A01 26-35 50-66  99-107 EGVAAGEDY(SEQ ID NO: 3123) I079A03 26-35 50-66  99-107 GGMDWDFDY(SEQ ID NO: 3183) I079A04 26-35 50-66  99-107 VDSSGYAYY(SEQ ID NO: 3213) I079A06 26-35 50-66  99-106 DAAVTAEG (SEQ ID NO: 3142)I079A07 26-35 50-66  99-109 GSNYSPDAFDI (SEQ ID NO: 3112) I079A10 26-3550-66 101-121 LPPDLRYCDGGICPGFDWLGP (SEQ ID NO: 3163) I079A11 26-3550-66  99-108 GPSYYYYMAV (SEQ ID NO: 3114) I079B02 26-35 50-66  99-107EGVAAGEDY (SEQ ID NO: 3123) I079B03 26-35 50-66  99-109 GSNYSPDAFDI(SEQ ID NO: 3112) I079B04 26-35 50-66  99-103 LLSDY (SEQ ID NO: 3168)I079B07 26-35 50-66  99-111 DLSGSYFSRYFDY (SEQ ID NO: 3193) I079B0926-35 50-66  99-112 VEWEDIVVGSAFDI (SEQ ID NO: 3128) I079C02 26-35 50-66 99-117 VTSLYSSSSGGYYYYGMDV (SEQ ID NO: 3145) I079C04 26-35 50-66 99-105 GWRGVDY (SEQ ID NO: 3195) I079C05 26-35 50-66  99-113AGGNPRSGSLVYFDY (SEQ ID NO: 3225) I079C07 26-35 50-66  99-110GLDVYAIYGLDV (SEQ ID NO: 3176) I079D01 26-35 50-66  99-117EVRNYDLLTRSYLAGPLDN (SEQ ID NO: 2751) I079D02 26-35 50-66  99-108EIGWEGAFDI (SEQ ID NO: 3178) I079D04 26-35 50-66  99-106 VRPGLMDV(SEQ ID NO: 3132) I079D06 26-35 50-66  99-110 EAYTSSWAEFDF(SEQ ID NO: 3190) I079D07 26-35 50-66  99-109 NITPLAMVGDF(SEQ ID NO: 3146) I079D08 26-35 50-66  99-103 LIEDF (SEQ ID NO: 3161)I079D09 26-35 50-66  99-104 DSGSPD (SEQ ID NO: 3108) I079D11 26-35 50-66 99-107 EGVAAGEDY (SEQ ID NO: 3123) I079E06 26-35 50-66  99-109EKRGSRRVFDI (SEQ ID NO: 3093) I079E08 26-35 50-66  99-110 EAYASSWAEFDF(SEQ ID NO: 3189) I079E11 26-35 50-66  99-109 PYGSGSYAFDI(SEQ ID NO: 3185) I079E12 26-35 50-66  99-116 ARDYYDSSGYYVPDAFDI(SEQ ID NO: 3107) I079F01 26-35 50-66  99-106 GHFYGMDV (SEQ ID NO: 3098)I079F02 26-35 50-68 101-121 LPPDLRYCDGGMCSGFDWLGP (SEQ ID NO: 3219)I079F03 26-35 50-66  99-113 ESLLTEEYCGSDCYS (SEQ ID NO: 3115) I079F0426-35 50-66  99-109 NSAPPAPSMDV (SEQ ID NO: 3099) I079F09 26-35 50-66 99-103 RYYDY (SEQ ID NO: 3139) I079F10 26-35 50-66  99-109 NITPLAMVGDF(SEQ ID NO: 3146) I079F12 26-35 50-66  99-109 ADYSNDYYMDV(SEQ ID NO: 3166) I079G02 26-35 50-66  99-109 NITPLAMVGDF(SEQ ID NO: 3146) I079G05 26-35 50-66  99-109 FPLESYYYMDV(SEQ ID NO: 3124) I079G06 26-35 50-66  99-108 GNSFGRTLDY(SEQ ID NO: 3158) I079H05 26-35 50-66  99-109 DVPPPDGYLEV(SEQ ID NO: 3192) I079H06 26-35 50-66  99-107 ASYPVPFDY(SEQ ID NO: 3171) I080A01 26-35 50-66  99-104 GGWLDD (SEQ ID NO: 3210)I080A02 26-35 50-66  99-106 EHSSSFDY (SEQ ID NO: 3111) I080A05 26-3550-66  99-114 EGEGDGYNVAPYYFDY (SEQ ID NO: 3160) I080A06 26-35 50-66 99-114 EAGGSGSYHFSFPFDY (SEQ ID NO: 3188) I080A07 26-35 50-66  99-108TGIWGYYFDY (SEQ ID NO: 3175) I080A10 26-35 50-66  99-114DGNLNYDGSTDYGMDV (SEQ ID NO: 3140) I080B02 26-35 50-66  99-111LGRNYTSSWSLDY (SEQ ID NO: 3181) I080B03 26-35 50-66  99-111VVGGYSSTLGTDV (SEQ ID NO: 3096) I080B05 26-35 50-66  99-110 LGVARGREAFDL(SEQ ID NO: 3206) I080B06 26-37 52-69 102-115 AVRSPGYYYYYMDV(SEQ ID NO: 3125) 1080B07 26-35 50-66  99-106 GRKPLFDY (SEQ ID NO: 3141)I080B08 26-37 52-67 100-109 KQRREKYFDY (SEQ ID NO: 3100) I080B09 26-3550-66  99-115 EKAIIETTSGEADPFDI (SEQ ID NO: 3151) I080B10 26-37 52-67100-111 RPALRSLWYFDL (SEQ ID NO: 3102) I080B11 26-35 50-68 101-110LHCTGGSCGF (SEQ ID NO: 3186) I080B12 26-35 50-66  99-112 NPYYYDSSEGFFDY(SEQ ID NO: 3109) I080003 26-35 50-66  99-111 SGRQAYYYYGMDV(SEQ ID NO: 3091) I080006 26-36 51-66  99-117 DYYDGSSYSSGDYYYYMDV(SEQ ID NO: 3227) I080007 26-35 50-66  99-117 DSDLVVIPTAIQGRYYFDN(SEQ ID NO: 3113) I080008 26-35 50-66  99-110 GKRYSYGWYFDI(SEQ ID NO: 3130) I080C10 26-35 50-66  99-104 DTPLDP (SEQ ID NO: 3094)I080C11 26-35 50-66  99-110 EGDPTDNDAFDV (SEQ ID NO: 3155) I080C12 26-3550-66  99-111 DGPTYARPYYLDH (SEQ ID NO: 3153) I080D01 26-35 50-66 99-109 DGTKYDWGFDY (SEQ ID NO: 3220) I080D02 26-35 50-66  99-114ETFSHCSGGSCYPFDY (SEQ ID NO: 3212) I080D04 26-35 50-66  99-111SGRQAYYYYGMDV (SEQ ID NO: 3091) I080D05 26-35 50-66  99-109 EFFGYVYLTDY(SEQ ID NO: 3165) I080D08 26-35 50-68 101-110 LHCTGGSCGF(SEQ ID NO: 3186) I080D09 26-35 50-66  99-111 VDYTDYEMGAFEI(SEQ ID NO: 3187) I080D11 26-35 50-66  99-108 VGNFGYYFEY(SEQ ID NO: 3196) I080D12 26-35 50-68 101-108 SSRNGGDY (SEQ ID NO: 3214)I080E01 26-35 50-66  99-109 DLSRVAGRFDY (SEQ ID NO: 3164) I080E04 26-3752-67 100-109 HDVYGDLFDY (SEQ ID NO: 3211) I080E06 26-35 50-68 101-110LHCSGGSCGF (SEQ ID NO: 3221) I080E07 26-35 50-66  99-115EGSIVGATLTINDAFDI (SEQ ID NO: 3150) I080E08 26-35 50-66  99-110GKRYSYGWYFDI (SEQ ID NO: 3130) I080E12 26-35 50-66  99-103 DPFDY(SEQ ID NO: 3134) I080F04 26-35 50-66  99-111 DGPTYARPYYLDH(SEQ ID NO: 3153) I080F05 26-35 50-66  99-115 ESSGTLGEFSLELPFDY(SEQ ID NO: 3203) I080F06 26-35 50-66  99-111 LGRNYTSSWSLDY(SEQ ID NO: 3181) I080F08 26-35 50-66  99-103 NAFDY (SEQ ID NO: 3121)I080G03 26-36 50-66  99-113 GRGYSSSSSVYGMDI (SEQ ID NO: 3095) I080G0426-35 50-66  99-104 VHSSGS (SEQ ID NO: 3216) I080G10 26-35 50-66  99-116KRGDFGVIRLHHYYGMDV (SEQ ID NO: 3136) I080G11 26-37 52-67 100-109HDVYGDLFDS (SEQ ID NO: 3205) I080H01 26-37 52-67 100-113 LRPDADYGDYGFDY(SEQ ID NO: 3218) I080H02 26-35 50-66  99-112 TSERGTYRQWDFDN(SEQ ID NO: 3204) I080H03 26-35 50-66  99-108 EAGEVAAIDY(SEQ ID NO: 3180) I080H04 26-35 50-66  99-110 GKRYSYGWYFDI(SEQ ID NO: 3130) I080H05 26-37 52-67 100-109 HDVYGDLFDS(SEQ ID NO: 3205) I080H06 26-35 50-66  99-110 GKRYSYGWYFDV(SEQ ID NO: 3217) I080H07 26-35 50-68 101-110 LHCTGGSCGF(SEQ ID NO: 3186) I080H08 26-35 50-66  99-111 ERGGRDGDYALFD(SEQ ID NO: 3148) I080H09 26-36 51-66  99-112 RTPDHNGDSGPPDY(SEQ ID NO: 3215) I081A01 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I081A03 26-35 50-66  99-108 ESLTGGAFDI (SEQ ID NO: 3117) I081A04 26-3550-66  99-103 DTTDY (SEQ ID NO: 2203) I081A06 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I081A08 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I081A09 26-35 50-66  99-107 GAGSRYFDL (SEQ ID NO: 3118) I081A10 26-3550-66  99-106 GGDRAFDI (SEQ ID NO: 3119) I081B01 26-35 50-66  99-103DTTDY (SEQ ID NO: 2203) I081B04 26-35 50-66  99-107 GNAWGAFDI(SEQ ID NO: 2211) I081B05 26-35 50-66  99-106 GGDRAFDI (SEQ ID NO: 3119)I081B06 26-35 50-66  99-106 VKRYYFDY (SEQ ID NO: 3179) I081B07 26-3550-66  99-109 ELTGANDAFDI (SEQ ID NO: 3104) I081B08 26-35 50-66  99-105RRYALDY (SEQ ID NO: 2920) I081B09 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I081B10 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I081B11 26-35 50-66  99-105 GFALYKD (SEQ ID NO: 3169) I081C07 26-3550-66  99-103 DTTDY (SEQ ID NO: 2203) I081C08 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I081D04 26-35 50-66  99-108 EDLTGDAFDI(SEQ ID NO: 3103) I081D06 26-35 50-66  99-105 GDAYFDY (SEQ ID NO: 3147)I081D08 26-35 50-66  99-105 GDAYFDY (SEQ ID NO: 3147) I081D09 26-3550-66  99-103 DTTDY (SEQ ID NO: 2203) I081D10 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I081D11 26-35 50-66  99-107 EGLLDAFDI(SEQ ID NO: 3200) I081D12 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I081E02 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203) I081E03 26-35 50-66 99-103 DTTDY (SEQ ID NO: 2203) I081E05 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I081E06 26-35 50-66  99-107 VGYGGKGDY(SEQ ID NO: 3137) I081E07 26-35 50-66  99-107 GAGSRYFDL(SEQ ID NO: 3118) I081E10 26-35 50-66  99-115 GLAPIVDGGMTNDAFDI(SEQ ID NO: 3184) I081F01 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I081F04 26-35 50-66  99-105 RLIRKAR (SEQ ID NO: 3170) I081F05 26-3550-66  99-103 DTTDY (SEQ ID NO: 2203) I081F06 26-35 50-66  99-107ERGNQAFDI (SEQ ID NO: 3156) I081F07 26-35 50-66  99-105 RRYALDY(SEQ ID NO: 2920) I081F11 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I081G01 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203) I081G04 26-35 50-66 99-103 DTTDY (SEQ ID NO: 2203) I081G06 26-35 50-66  99-108 SRSPYDAFDI(SEQ ID NO: 3097) I081G10 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I081H02 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203) I081H03 26-35 50-66 99-103 DTTDY (SEQ ID NO: 2203) I081H04 26-35 50-66  99-108 SNWGGDAFDI(SEQ ID NO: 3202) I081H06 26-35 50-66  99-103 LAFDI (SEQ ID NO: 3174)I081H08 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203) I082A02 26-35 50-66 99-112 PAASSRGPKDAFDI (SEQ ID NO: 3129) I082A04 26-35 50-66  99-103LSGDS (SEQ ID NO: 3122) I082A08 26-35 50-66  99-107 EGVAAGEDY(SEQ ID NO: 3123) I082A11 26-35 50-66  99-103 FVLDY (SEQ ID NO: 2210)I082B06 26-35 50-66  99-104 GNGKDV (SEQ ID NO: 3135) I082B09 26-35 50-66 99-107 EGVAAGEDY (SEQ ID NO: 3123) I082B12 26-35 50-66  99-104 DLDFDY(SEQ ID NO: 2208) I082C01 26-35 50-66  99-109 VNDIVVVDMDV(SEQ ID NO: 3143) I082C05 26-35 50-66  99-109 EKRGSRRVFDI(SEQ ID NO: 3093) I082C08 26-35 50-66  99-110 LSNRNDNLRLDY(SEQ ID NO: 3106) I082D02 26-35 50-66  99-103 FVLDY (SEQ ID NO: 2210)I082E05 26-35 50-66  99-107 TWATNTFDM (SEQ ID NO: 3152) I082E06 26-3550-66  99-103 FDLDY (SEQ ID NO: 3167) I082E07 26-35 50-66  99-112VEWEDIVVGSAFDI (SEQ ID NO: 3128) I082F11 26-35 50-66  99-109 GGDMTTVTTDY(SEQ ID NO: 3177) I082G07 26-35 50-66  99-109 ADYSNDYYMDV(SEQ ID NO: 3166) I082G10 26-35 50-66  99-107 EGVAAGEDY(SEQ ID NO: 3123) I082G11 26-35 50-66  99-116 GPIYYFDGSAYEGYYFDY(SEQ ID NO: 3222) I082H04 26-35 50-65  99-105 MNADAFEI (SEQ ID NO: 3223)I082H09 26-35 50-66  99-112 PAASSRGPKDAFDI (SEQ ID NO: 3129) I083A0626-35 50-66  99-109 DSRPTNRAFDY (SEQ ID NO: 3110) I083A09 26-35 50-68101-110 LHCTGGSCGF (SEQ ID NO: 3186) I083A11 26-35 50-66  99-108VRDDSAGFDY (SEQ ID NO: 3173) I083B03 26-35 50-66  99-110 VLVRGQYRGMDL(SEQ ID NO: 3138) I083B05 26-35 50-66  99-111 VDYTDYEMGAFDL(SEQ ID NO: 3172) I083B06 26-35 50-66  99-111 DRIAAAGGDAFDI(SEQ ID NO: 3194) I083B10 26-35 50-66  99-110 DLYKNGYALFDS(SEQ ID NO: 3197) I083C01 26-35 50-66  99-108 DEYSSLYMDV(SEQ ID NO: 3201) I083C02 26-35 50-66  99-108 FGAGRLYDDY(SEQ ID NO: 3224) I083C07 26-35 50-66  99-109 DNGGGTIGFDY(SEQ ID NO: 2195) I083C12 26-35 50-66  99-108 DQGIETANDY(SEQ ID NO: 3207) I083D04 26-35 50-66  99-118 DILPDYDFWNPNEDASSLDT(SEQ ID NO: 3133) I083D07 26-35 50-66  99-121 DFQMVRGVFIANPPIYNYYGMDV(SEQ ID NO: 3154) I083D08 26-35 50-66  99-115 DADEGLVEAETTNWFDS(SEQ ID NO: 3126) I083D10 26-37 52-69 102-119 ATKSYDILTRMYYYHMDV(SEQ ID NO: 2748) I083D12 26-35 50-66  99-105 DRTRMDV (SEQ ID NO: 3182)I083E02 26-35 50-66  99-111 VGIKAAAVDNFEY (SEQ ID NO: 2197) I083E0326-35 50-66  99-108 DEIYNDAFDY (SEQ ID NO: 3105) I083E04 26-35 50-66 99-116 DGDISDSPINNWNYAMDI (SEQ ID NO: 3101) I083E08 26-35 50-66  99-111RGGTSENYSGMDV (SEQ ID NO: 3209) I083E12 26-35 50-66  99-107 DYPHNAFDI(SEQ ID NO: 3127) I083F02 26-35 50-66  99-118 DVRSDRFWSGGYFHYSGMDV(SEQ ID NO: 3131) I083F04 26-35 50-66  99-110 STLEVGATDFDY(SEQ ID NO: 3199) I083F06 26-35 50-66  99-107 SDDWGAYHI(SEQ ID NO: 3198) I083F08 26-35 50-66  99-111 ERGGRDGDYALDF(SEQ ID NO: 3148) I083F11 26-35 50-66  99-109 ELVGAPGGFDP(SEQ ID NO: 3191) I083G04 26-35 50-68  99-111 VDYTDYEMGAFDL(SEQ ID NO: 3172) I083G05 26-35 50-66 101-110 SVAGRGNFDY(SEQ ID NO: 3208) I083G06 26-35 50-66  99-111 ERGGRDGDYALDF(SEQ ID NO: 3148) I083G08 26-35 50-66  99-114 EGGGDAYDVAPYYFDY(SEQ ID NO: 2204) I083G09 26-35 50-66  99-103 DPFDY (SEQ ID NO: 3134)I083G11 26-35 50-66  99-113 ALLGLPSDFSYYVDV (SEQ ID NO: 3159) I083H0426-35 50-66  99-114 EGEGDGYNVAPYYFDY (SEQ ID NO: 3160) I083H05 26-3550-66  99-106 TDYGGFDY (SEQ ID NO: 3092) I083H07 26-35 50-66  99-110GGVGDSRGVFDP (SEQ ID NO: 3162) I084A03 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I084A08 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I084B08 26-35 50-66  99-108 ESLTGDAFDI (SEQ ID NO: 3116) I084C02 26-3550-66  99-109 SPLHFSDAFDI (SEQ ID NO: 3120) I084D03 26-35 50-66  99-103DTTDY (SEQ ID NO: 2203) I084D05 26-35 50-66  99-106 EVGGAFDI(SEQ ID NO: 3157) I084E01 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I084E06 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203) I084E10 26-35 50-66 99-103 DTTDY (SEQ ID NO: 2203) I084E12 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I084F04 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I084F07 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203) I084F12 26-35 50-66 99-108 ESLTGDAFDI (SEQ ID NO: 3116) I084G12 26-35 50-66  99-103 DTTDY(SEQ ID NO: 2203) I084H02 26-35 50-66  99-103 DTTDY (SEQ ID NO: 2203)I099B05 26-35 50-66  99-118 GAHYYDRSPSHLKSYWYFDL (SEQ ID NO: 3149)I099G09 26-35 50-66  99-111 VGIKAAAVDNFEY (SEQ ID NO: 2197) I099H0126-35 50-66  99-111 LGRNYTSSWSLDY (SEQ ID NO: 3181) I099H06 26-35 50-66 99-111 VGIKAAAVDNFEY (SEQ ID NO: 2197) I099H08 26-35 50-66  99-117GGRYGYYYDGTGYVDAFDI (SEQ ID NO: 3226) I100A01 26-35 50-66  99-109DNGGGTIGFDY (SEQ ID NO: 2195) I100A10 26-35 50-66  99-113VRQQIADPPRSFFDP (SEQ ID NO: 3144) I100B03 26-35 50-66  99-109DNGGGTIGFDY (SEQ ID NO: 2195) I100B04 26-35 50-66  99-109 DNGGGTIGFDY(SEQ ID NO: 2195) I100003 26-35 50-66  99-113 VRQQIADPPRSFFDP(SEQ ID NO: 3144) BAB2001 31-35 50-65  99-113 EQYDTLTGSPYGMDV BAB208031-35 50-66  99-108 LNSLRGGHDY BAB2015 31-35 50-65  99-113GASSGWYDYYYYMDV BAB2019 31-35 50-66  99-114 DSYDILTDYYNMIMDV BAB208731-35 50-66  99-117 GFTGYDILTDYYSVDYFDS BAB2016 31-35 50-66  99-114DPRYDILTGYLYGMDV BAB2034 31-37 52-69 102-121 EGAHYDILTGHNYYHYGMDVBAB2065 31-35 50-66  99-114 ATYDPLTGYSFDGFD

1. A method of treating an individual clinically diagnosed with anautoimmune disease, comprising: analyzing a biological sample from anindividual clinically diagnosed with autoimmune disease for the presenceor absence of elevated BCMA protein expression levels on their B cells,wherein the presence of elevated BCMA protein expression levels isassociated with the clinical diagnosis of autoimmune disease; andselecting a treatment plan that is most effective for individualsclinically diagnosed as having a condition associated with an increasedBCMA protein expression level.
 2. The method of claim 1 wherein saidtreatment plan involves administration of a BLyS antagonist.
 3. Themethod of claim 2 wherein said BLyS antagonist is also an APRILantagonist.
 4. The method of claim 1, wherein the autoimmune disease isselected from the group consisting of rheumatoid arthritis, juvenilerheumatoid arthritis, systemic lupus erythematosus (SLE), lupusnephritis (LN), Wegener's disease, inflammatory bowel disease,idiopathic thrombocytopenic purpura (ITP), thrombotic throbocytopenicpurpura (TTP), autoimmune thrombocytopenia, multiple sclerosis,psoriasis, IgA nephropathy, IgM polyneuropathies, myasthenia gravis,vasculitis, diabetes mellitus, Reynaud's syndrome, Sjorgen's syndromeand glomerulonephritis.
 5. The method of claim 1 wherein said autoimmunedisease is SLE.
 6. A method for predicting a patient's likelihood torespond to a drug treatment for an autoimmune disease, comprisingdetermining the level of BCMA protein expression on the patient's Bcells, wherein the presence of elevated BCMA protein expression levelsis predictive of the patient's likelihood to respond to a drug treatmentfor the condition.
 7. The method of claim 6 wherein the autoimmunedisease is selected from the group consisting of rheumatoid arthritis,juvenile rheumatoid arthritis, systemic lupus erythematosus (SLE), lupusnephritis (LN), Wegener's disease, inflammatory bowel disease,idiopathic thrombocytopenic purpura (ITP), thrombotic throbocytopenicpurpura (TTP), autoimmune thrombocytopenia, multiple sclerosis,psoriasis, IgA nephropathy, IgM polyneuropathies, myasthenia gravis,vasculitis, diabetes mellitus, Reynaud's syndrome, Sjorgen's syndromeand glomerulonephritis.
 8. The method of claim 6 wherein said autoimmunedisease is SLE.
 9. The method of claim 6 wherein said drug treatmentinvolves administration of a BLyS antagonist.
 10. The method of claim 9wherein said BLyS antagonist is also an APRIL antagonist.
 11. An invitro method of selecting a treatment plan that is most effective fortreating an individual clinically diagnosed with an autoimmune disease,comprising: analyzing in vitro a biological sample from an individualclinically diagnosed with autoimmune disease for the presence or absenceof elevated BCMA levels on their B cells, wherein the presence ofelevated BCMA levels is associated with the clinical diagnosis ofautoimmune disease.
 12. The method of claim 11 wherein said treatmentplan involves the use of a BLyS antagonist.
 13. The method of claim 12wherein said BLyS antagonist is also an APRIL antagonist.
 14. The methodof claim 11, wherein the autoimmune disease is selected from the groupconsisting of rheumatoid arthritis, juvenile rheumatoid arthritis,systemic lupus erythematosus (SLE), lupus nephritis (LN), Wegener'sdisease, inflammatory bowel disease, idiopathic thrombocytopenic purpura(ITP), thrombotic throbocytopenic purpura (TTP), autoimmunethrombocytopenia, multiple sclerosis, psoriasis, IgA nephropathy, IgMpolyneuropathies, myasthenia gravis, vasculitis, diabetes mellitus,Reynaud's syndrome, Sjorgen's syndrome and glomerulonephritis.
 15. Themethod of claim 11 wherein said autoimmune disease is SLE.
 16. An invitro method for predicting a patient's likelihood to respond to a drugtreatment for an autoimmune disease, comprising determining the level ofBCMA expression on the surface of B cells in a sample from the patient;wherein the presence of elevated B cell levels is predictive of thepatient's likelihood to respond to a drug treatment for the condition.17. The method of claim 16 wherein the autoimmune disease is selectedfrom the group consisting of rheumatoid arthritis, juvenile rheumatoidarthritis, systemic lupus erythematosus (SLE), lupus nephritis (LN),Wegener's disease, inflammatory bowel disease, idiopathicthrombocytopenic purpura (ITP), thrombotic throbocytopenic purpura(TTP), autoimmune thrombocytopenia, multiple sclerosis, psoriasis, IgAnephropathy, IgM polyneuropathies, myasthenia gravis, vasculitis,diabetes mellitus, Reynaud's syndrome, Sjorgen's syndrome andglomerulonephritis.
 18. The method of claim 16 wherein said autoimmunedisease is SLE.
 19. The method of claim 16 wherein said drug treatmentcomprises a BLyS antagonist.
 20. The method of claim 19 wherein saidBLyS antagonist is also an APRIL antagonist.
 21. A BLyS antagonist foruse in the treatment of an autoimmune disease in a patient, wherein saidpatient has elevated levels of BCMA expression on the surface of Bcells.
 22. The antagonist of claim 21 wherein the autoimmune disease isSLE.
 23. The antagonist of claim 21 wherein said antagonist is a BLySantibody.
 24. The antagonist of claim 23 wherein said BLyS antibody isLymphostat-B.
 25. The antagonist of claim 23 wherein said antagonist isa receptor-extracellular domain/Fc domain fusion protein selected fromthe group consisting of TACI-Ig, BCMA-Ig, and BAFF-R-Ig.
 26. Theantagonist of claim 25 wherein said receptor-extracellular domain/Fcdomain fusion protein is TACI-Ig.
 27. The antagonist of claim 26 whereinsaid TACI-Ig is atacicept.
 28. A method of treating an individualdiagnosed with an autoimmune disease, comprising: analyzing a biologicalsample from an individual diagnosed with an autoimmune disease for thepresence or absence of elevated B Lymphocyte Stimulator receptor proteinexpression levels, wherein the presence of elevated B LymphocyteStimulator receptor protein expression levels is associated with thediagnosis of autoimmune disease; and treating the individual.
 29. Themethod of claim 28, wherein the method further comprises selecting atreatment plan that is most effective for individuals diagnosed ashaving a condition associated with an increased B Lymphocyte Stimulatorreceptor protein expression level prior to treating the individual 30.The method of claim 28, wherein the B Lymphocyte Stimulator receptor isBCMA.
 31. The method of claim 29, wherein the B Lymphocyte Stimulatorreceptor is BCMA.
 32. The method of claim 28, wherein treating theindividual comprises administering a BLyS antagonist.
 33. The method ofclaim 29, wherein the treatment plan involves administration of a BLySantagonist.
 34. The method of claim 28, wherein the BLyS antagonist isalso an APRIL antagonist.
 35. The method of claim 29, wherein the BLySantagonist is also an APRIL antagonist.
 36. The method of claim 28,wherein the autoimmune disease is selected from the group consisting ofrheumatoid arthritis, systemic lupus erythematosus (SLE), nephritisassociated with systemic lupus erythematosus, inflammatory boweldisease, idiopathic thrombocytopenic purpura (ITP), autoimmunethrobocytopenic purpura, autoimmune thrombocytopenia, multiplesclerosis, psoriasis, IgA nephropathy, IgM polyneuropathies, myastheniagravis, vasculitis, diabetes mellitus, Reynaud's syndrome, Sjorgen'ssyndrome and glomerulonephritis.
 37. The method of claim 29, wherein theautoimmune disease is selected from the group consisting of rheumatoidarthritis, systemic lupus erythematosus (SLE), nephritis associated withsystemic lupus erythematosus, inflammatory bowel disease, idiopathicthrombocytopenic purpura (ITP), autoimmune throbocytopenic purpura,autoimmune thrombocytopenia, multiple sclerosis, psoriasis, IgAnephropathy, IgM polyneuropathies, myasthenia gravis, vasculitis,diabetes mellitus, Reynaud's syndrome, Sjorgen's syndrome andglomerulonephritis.
 38. The method of claim 28, wherein the autoimmunedisease is SLE.
 39. The method of claim 29, wherein the autoimmunedisease is SLE.
 40. A method for determining whether or not a patientdiagnosed with an autoimmune disease will respond to treatment with aBLyS antagonist, comprising determining the level of B LymphocyteStimulator receptor protein expression on the patient's cells, whereinthe presence of elevated B Lymphocyte Stimulator receptor proteinexpression levels is indicative of whether or not the patient willrespond to the treatment.
 41. The method of claim 40, wherein the BLymphocyte Stimulator receptor is BCMA.
 42. The method of claim 40,wherein the autoimmune disease is selected from the group consisting ofrheumatoid arthritis, systemic lupus erythematosus (SLE), nephritisassociated with systemic lupus erythematosus, inflammatory boweldisease, idiopathic thrombocytopenic purpura (ITP), autoimmunethrombocytopenic purpura, autoimmune thrombocytopenia, multiplesclerosis, psoriasis, IgA nephropathy, IgM polyneuropathies, myastheniagravis, vasculitis, diabetes mellitus, Reynaud's syndrome, Sjorgen'ssyndrome and glomerulonephritis.
 43. The method of claim 40, wherein theautoimmune disease is SLE.
 44. The method of claim 40, wherein the BLySantagonist is also an APRIL antagonist.
 45. An in vitro method ofselecting a treatment that is most effective for treating an individualdiagnosed with an autoimmune disease, comprising analyzing in vitro abiological sample from an individual diagnosed with an autoimmunedisease for the presence or absence of elevated B Lymphocyte Stimulatorreceptor levels on their cells, wherein the presence of elevated BLymphocyte Stimulator receptor levels is associated with the diagnosisof the autoimmune disease.
 46. The method of claim 45, wherein the BLymphocyte Stimulator receptor is BCMA.
 47. The method of claim 45,wherein the treatment involves the use of a BLyS antagonist.
 48. Themethod of claim 47, wherein the BLyS antagonist is also an APRILantagonist.
 49. The method of claim 45, wherein the autoimmune diseaseis selected from the group consisting of rheumatoid arthritis, systemiclupus erythematosus (SLE), nephritis associated with systemic lupuserythematosus, inflammatory bowel disease, idiopathic thrombocytopenicpurpura (ITP), autoimmune thrombocytopenic purpura, autoimmunethrombocytopenia, multiple sclerosis, psoriasis, IgA nephropathy, IgMpolyneuropathies, myasthenia gravis, vasculitis, diabetes mellitus,Reynaud's syndrome, Sjorgen's syndrome and glomerulonephritis.
 50. Themethod of claim 45, wherein the autoimmune disease is SLE.
 51. An invitro method for determining whether or not a patient diagnosed with anautoimmune disease will respond to treatment with a BLyS antagonist,comprising determining the level of B Lymphocyte Stimulator receptorexpression on the surface of cells in a sample from the patient; whereinthe presence of elevated B Lymphocyte Stimulator receptor levels isindicative of whether or not the patient will respond to the treatment.52. The method of claim 51, wherein the B Lymphocyte Stimulator receptoris BCMA.
 53. The method of claim 51, wherein the autoimmune disease isselected from the group consisting of rheumatoid arthritis, systemiclupus erythematosus (SLE), nephritis associated with systemic lupuserythematosus, inflammatory bowel disease, idiopathic thrombocytopenicpurpura (ITP), autoimmune thrombocytopenic purpura, autoimmunethrombocytopenia, multiple sclerosis, psoriasis, IgA nephropathy, IgMpolyneuropathies, myasthenia gravis, vasculitis, diabetes mellitus,Reynaud's syndrome, Sjorgen's syndrome and glomerulonephritis.
 54. Themethod of claim 51, wherein the autoimmune disease is SLE.
 55. Themethod of claim 51, wherein the BLyS antagonist is also an APRILantagonist.
 56. A BLyS antagonist for use in the treatment of anautoimmune disease in a patient, wherein the patient has elevated levelsof B Lymphocyte Stimulator receptor expression on the surface of cells.57. The antagonist of claim 56, wherein the B Lymphocyte Stimulatorreceptor is BCMA.
 58. The antagonist of claim 56, wherein the autoimmunedisease is SLE.
 59. The antagonist of claim 56, wherein the antagonistis a BLyS antibody.
 60. The antagonist of claim 59, wherein the BLySantibody comprises amino acid residues 1-123 of SEQ ID NO: 327 and aminoacid residues 141-249 of SEQ ID NO:
 327. 61. The antagonist of claim 56,wherein the antagonist is a receptor-extracellular domain/Fc domainfusion protein selected from the group consisting of TACI-Ig, BCMA-Ig,and BAFF-R-Ig.
 62. The antagonist of claim 56, wherein thereceptor-extracellular domain/Fc domain fusion protein is TACI-Ig.